Biophotonic compositions and methods for providing biophotonic treatment

ABSTRACT

The present disclosure provides biophotonic topical compositions and methods useful in phototherapy. In particular, the biophotonic topical compositions of the present disclosure are substantially resistant to leaching such that very low amounts of chromophore(s) present in the biophotonic composition leach out of the composition. The biophotonic compositions and the methods of the present disclosure are useful for promoting wound healing and skin rejuvenation, as well as treating acne and various skin disorders.

CROSS-REFERENCE TO RELATED APPLICATION

This application claims the benefit of U.S. Provisional Application Nos.61/636,480, filed Apr. 20, 2012; 61/701,502, filed Sep. 14, 2012;61/636.574 filed on Apr. 20, 2012; 61/701,510, filed on Sep. 14, 2012;61/636,577, filed on Apr. 20, 2012: 61/701,513, filed on Sep. 14, 2012;and 61/766,611, filed on Feb. 19, 2013; which applications are herebyincorporated by reference in their entireties.

BACKGROUND OF THE DISCLOSURE

Phototherapy has recently been recognized as having wide range ofapplications in both the medical, cosmetic and dental fields for use insurgeries, therapies and examinations. For example, phototherapy hasbeen developed to treat cancers and tumors with lessened invasiveness.Phototherapy has also been used to disinfect target sites as anantimicrobial treatment. Phototherapy has also been found to promotewound healing.

Photodynamic therapy is a type of phototherapy which involves a step ofsystemic administration or uptake of a photosensitive agent into thediseased or injured tissue, which step is followed by site-specificapplication of activating light (photodynamic therapy). Such regimens,however, are often associated with undesired side-effects, includingsystemic or localized toxicity due to the direct contact of thephotosensitive agents with the tissues. Moreover, such existing regimensoften demonstrate low therapeutic efficacy due to, for example, the pooruptake of the photosensitive agents into the target tissues. Therefore,it is an object of the present disclosure to provide new and improvedcompositions and methods useful in phototherapy.

SUMMARY OF THE DISCLOSURE

The present disclosure provides topical biophotonic compositions andmethods useful in phototherapy. In particular, the biophotoniccompositions of the present disclosure may contain a gelling agent thatprovides a barrier such that the chromophore(s) or photosensitiveagent(s) and other components of the topical biophotonic compositionsare not in substantial contact with the target tissues, and/or do notpenetrate the target tissues. Put another way, the biophotoniccompositions of the present disclosure may contain a gelling agent,which provides a barrier rendering the compositions substantiallyresistant to leaching. The use of such biophotonic topical compositionsm phototherapy would therefore not involve substantial direct contact ofthe target tissues with a photosensitizing agent or chromophore, whichmay be potentially toxic to or may cause undesired side effects at thetissues.

In some aspects, there is provided a topical biophotonic compositioncomprising at least a first chromophore and a gelling agent, wherein thebiophotonic composition is substantially resistant to leaching such thatless than 15% of the total chromophore amount leaches out of thebiophotonic composition into tissue. In some embodiments, thebiophotonic composition is substantially resistant to leaching such thatless than 15% of total chromophore amount leaches out of the biophotoniccomposition into tissue when in contact with the tissue for at leastabout, 5 minutes, about 10 minutes, about 15 minutes, about 20 minutes,about 25 minutes or about 30 minutes. In some embodiments, less than 15%of total chromophore amount leaches out of the biophotonic compositionduring a treatment time. The treatment time can be up to about 5minutes, about 10 minutes, about 15 minutes, about 20 minutes, about 25minutes, about 30 minutes.

In some aspects, there is provided a topical biophotonic compositioncomprising at least a first chromophore and a gelling agent, wherein thefirst chromophore is photoactive in the composition, and wherein thecomposition is substantially resistant to leaching such that less than15% of total chromophore amount can leach out into tissue during atreatment time in which the composition is topically applied ontotissue. The treatment time may comprise the total length of time thatthe composition is in contact with tissues, or if different, the time oflight illumination of the composition.

In some aspects, there is provided a topical biophotonic compositioncomprising at least a first chromophore and a gelling agent, wherein thebiophotonic composition is a gel or a semi-solid and is substantiallyresistant to leaching such that less than 15% of the total chromophoreamount leaches out of the biophotonic composition into tissue when incontact with tissue for at least about 5 minutes.

In some aspects, there is provided a topical biophotonic compositioncomprising at least a first chromophore and a gelling agent, wherein thebiophotonic composition is substantially translucent and issubstantially resistant to leaching such that less than 15% of the totalchromophore amount leaches out of the biophotonic composition intotissue when in contact with tissue for at least about 5 minutes. Bysubstantially translucent is meant having a transmission of more thanabout 20%.

In another aspect, there is provided a biophotonic compositioncomprising a first chromophore, wherein the composition is encapsulatedin a membrane which limits leaching of the first chromophore such thatless than 15% of the total chromophore amount leaches out into tissuewhen in contact with the tissue for at least about 5 minutes. Thebiophotonic composition may also comprise a carrier medium which may bea liquid, a gel or a semi-solid.

In certain embodiments of any of the foregoing or following, thebiophotonic topical composition allows less than 30%, 25%, 20%, 15%,10%, 5%, 19%, 0.8%, 0.5% or 0.1%, or essentially none of saidchromophore content to leach out of the biophotonic composition.

In certain embodiments of any of the foregoing or following, thebiophotonic topical composition further comprises a second chromophore.In certain embodiments of any of the foregoing or following, the firstchromophore of the biophotonic topical composition has an emissionspectrum that overlaps at least 5%, 10%, 20%, 25%, 30%, 40%, 50%, 60%,70% with an absorption spectrum of the second chromophore, when present.In some embodiments, the first chromophore of the biophotonic topicalcomposition has an emission spectrum that overlaps at least 1-10%,5-15%, 10-20%, 15-25%, 20-30%, 25-35%, 30-40%, 35-45%, 50-60%, 55-65% or60-70% with an absorption spectrum of the second chromophore whenpresent.

In certain embodiments of any of the foregoing or following, the gellingagent comprises a hygroscopic substance. In addition or m thealternative, the gelling agent may also be a hydrophilic polymer, ahydrated polymer or a lipid. In certain embodiments, the gelling agentcomprises one or more of glycerin, glycols such as propylene glycol,polyacrylic acid polymers, hyaluronic acid, glucosamine sulphate orgelatin.

In certain embodiments of any of the foregoing or following, the gellingagent is a high molecular weight, cross-linked polyacrylic acid polymerhaving a viscosity in the range of about 20,000-80,000, 20,000-100,000,25,000-90,000, 30,000-80,000, 30,000-70,000 30,000-60,000, 25,000-40,000cP. In certain embodiments, the cross-linked polyacrylic acid polymer isa carbomer selected from the group consisting of, but not limited to,Carbopol® 71G NF, 971P NF, 974P NF, 980 NF, 981 NF, 5984 EP, ETD 2020NF,Ultrez 10 NF, 934 NF, 934P NF, 940 NF, 941 NF, or 1342 NF.

In certain embodiments of any of the foregoing or following, thebiophotonic composition is substantially translucent and/or transparent.In certain embodiment, the biophotonic composition has a translucency ofat least 70% at 460 nm. In other embodiments, the composition has atranslucency of at least 20%, 30%, 40%, 50%, 60%, 70%, 75%, 85%, 90%,95% or 100% at 460 nm.

In certain embodiments of any of the foregoing or following, thebiophotonic composition is a gel or a semi-solid.

In certain embodiments of any of the foregoing or following, thebiophotonic composition is encapsulated in a transparent, impermeablemembrane, or a breathable membrane which allows permeation of gases butnot liquids. The membrane may comprise a lipid.

In certain embodiments of any of the foregoing or following, thebiophotonic composition further comprises an oxygen-generating agent. Insome embodiments, the oxygen-generating agent comprises hydrogenperoxide, carbamide peroxide, benzoyl peroxide, or water.

In certain embodiments of any of the foregoing or following, at leastone of the chromophores, for example, the first chromophore,photobleaches during illumination with light. In certain embodiments, atleast one of the chromophores, for example, the first chromophore emitsfluorescence upon illumination with light.

In certain embodiments of any of the foregoing or following,illumination of the biophotonic topical composition with light causes atransfer of energy from the first chromophore to the second chromophore.In some embodiments, the second chromophore emits fluorescence and/orgenerates reactive oxygen species after absorbing energy from the firstchromophore.

In certain embodiments of any of the foregoing or following, thebiophotonic composition does not generate a substantial amount of heatfollowing illumination with light. In some embodiments, the energyemitted by the biophotonic composition does not cause tissue damage.

In certain embodiments of any of the foregoing or following, the firstchromophore of the biophotonic topical composition absorbs light at awavelength of 200-600 nm, or 400-800 nm.

In certain embodiments of any of the foregoing or following, the firstchromophore absorbs light at a wavelength in the range of the visiblespectrum.

In certain embodiments of any of the foregoing or following, thebiophotonic composition comprises a second chromophore, which absorbslight at a wavelength in the range of the visible spectrum. In someembodiments, the second chromophore has an absorption wavelength that isrelatively longer than that of the first chromophore, for example,10-100 nm, 20-80 nm, 25-70 nm, or 30-60 nm longer.

In certain embodiments of any of the foregoing or following, the firstchromophore of the biophotonic topical composition is present in anamount of 0.01-40% per weight of the composition, and the secondchromophore, when present, is present in an amount of 0.001-40% perweight of the composition. In certain embodiments, the total weight perweight of chromophore or combination of chromophores may be in theamount of about 0.001-40.05% per weight of the composition.

In certain embodiments of any of the foregoing or following, thebiophotonic composition may be applied to or impregnated into a materialsuch as a pad, a dressing, a woven or non-woven fabric or the like. Theimpregnated material may be used as a mask (e.g. a face mask) or adressing.

In certain embodiments of any of the foregoing or following, thebiophotonic composition further comprises at least one waveguide withinor adjacent to the composition. The waveguide can be a particle, a fibreor a fibrillar network made of a material which can transmit and/or emitlight.

In certain embodiments of any of the foregoing or following, thecomposition does not comprise silica.

In certain embodiments of any of the foregoing or following, the firstor second chromophore is a fluorescent chromophore (‘fluorophore’).

In certain embodiments of any of the foregoing or following, the firstor second chromophore is a fluorescent xanthene. In some embodiments,the first or second chromophore is selected from Eosin Y, Erythrosin B,Fluorescein, Rose Bengal and Phloxin B. In certain embodiments, thebiophotonic composition comprises at least two of Eosin Y, Erythrosin B,Fluorescein, Rose Bengal and Phloxin B.

In certain embodiments of any of the foregoing or following, the firstchromophore is Eosin Y. In other embodiments, the first chromophore isFluorescein. In other embodiments, the first chromophore is Rose Bengal.In some embodiments, the biophotonic composition comprises Eosin andFluorescein. In other embodiments, the biophotonic composition comprisesEosin and Rose Bengal. In other embodiments, the biophotonic compositioncomprises Fluorescein and Rose Bengal. In other embodiments, thebiophotonic composition comprises Fluorescein and Rose Bengal.

In another aspect, there is provided a method for providing biophotonictherapy to a wound, comprising: applying a biophotonic composition to awound, wherein the biophotonic composition comprises at least at least afirst chromophore and a gelling agent; and illuminating the biophotoniccomposition with light having a wavelength that overlaps with anabsorption spectrum of the first chromophore: wherein the gelling agentrenders the biophotomic composition substantially resistant to leachingsuch that less than 15% of the total chromophore amount leaches out ofthe composition into tissue. In certain embodiments, the composition issubstantially resistant to leaching such that less than 15% of the totalchromophore amount leaches out of the biophotonic composition intotissue during a treatment time in which the composition is topicallyapplied onto tissue. In some embodiments, the biophotonic issubstantially resistant to leaching such that less than about 15% oftotal chromophore content leaches out of the biophotonic compositionduring a treatment time of at least 5 minutes, at least 10 minutes, atleast 15 minutes, at least 20 minutes, at least 25 minutes or at least30 minutes.

In some embodiments of the method for providing biophotonic therapy to awound, the method promotes wound healing. In certain embodiments of themethod, the wound as described herein includes for example chronic oracute wounds, such as diabetic foot ulcers, pressure ulcers, venousulcers or amputations. In some embodiments of the method for providingbiophotonic therapy to a wound, the method promotes reduction of scartissue formation.

In yet another aspect, there is provided a method for biophotonictreatment of acne comprising: applying a biophotonic composition to atarget skin tissue, wherein the biophotonic composition comprises atleast a first chromophore and a gelling agent; and illuminating saidbiophotonic composition with light having a wavelength that overlapswith an absorption spectrum of the first chromophore; wherein thegelling agent renders the biophotonic composition substantiallyresistant to leaching such that less than 15% of the total chromophoreamount leaches out of the composition into tissue. In certainembodiments, the composition is substantially resistant to leaching suchthat less than 15% of the total chromophore amount leaches out of thebiophotonic composition into tissue during a treatment time in which thecomposition is topically applied onto tissue. In some embodiments, thebiophotonic is substantially resistant to leaching such that less thanabout 15% of total chromophore content leaches out of the biophotoniccomposition during a treatment time of at least 5 minutes, at least 10minutes, at least 15 minutes, at least 20 minutes, at least 25 minutesor at least 30 minutes.

In certain embodiments of the method for biophotonic treatment acne, thetreatment can be applied to the skin tissue, such as on the face, once,twice, three times, four times, five times or six times a week, daily,or at any other frequency. The total treatment time can be one week, twoweeks, three weeks, four weeks, five weeks, six weeks, seven weeks,eight weeks, nine weeks, ten weeks, eleven weeks, twelve weeks, or anyother length of time deemed appropriate. In certain embodiments, theface may be split into separate areas (cheeks, forehead), and each areatreated separately. For example, the composition may be appliedtopically to a first portion, and that portion illuminated with light,and the biophotonic composition then removed. Then the composition isapplied to a second portion, illuminated and removed. Finally, thecomposition is applied to a third portion, illuminated and removed.

In certain embodiments of the method for biophotonic treatment ofwounds, the treatment can be applied in or on the wound once, twice,three times, four times, five times of six times a week, daily, or atany other frequency. The total treatment time can be one week, twoweeks, three weeks, four weeks, five weeks, six weeks, seven weeks,eight weeks, nine weeks, ten weeks, eleven weeks, twelve weeks, or anyother length of time deemed appropriate.

The disclosed methods for treating acne or wounds may further include,for example, administering a systemic or topical drug before, during orafter the biophotonic treatment. The drug may be an antibiotic, ahormone treatment, or any other pharmaceutical preparation which mayhelp to treat acne or wounds. The combination of a systemic treatmenttogether with a topical biophotonic treatment can reduce the duration ofsystemic treatment time.

In yet another aspect, there is provided a method for biophotonictreatment of a skin disorder comprising: applying a biophotoniccomposition to a target skin tissue, wherein the biophotonic compositioncomprises at least first chromophore and a gelling agent; andilluminating said biophotonic composition with light having a wavelengththat overlaps with an absorption spectrum of the first chromophore;wherein the gelling agent renders the biophotonic compositionsubstantially resistant to leaching such that less than 15% of the totalchromophore amount leaches out of the composition into tissue. Incertain embodiments, the composition as substantially resistant toleaching such that less than 15% of the total chromophore amount leachesout of the biophotonic composition into tissue during a treatment timein which the composition is topically applied onto tissue. In someembodiments, the biophotonic is substantially resistant to leaching suchthat less than about 15% of total chromophore content leaches out of thebiophotonic composition during a treatment time of at least 5 minutes,at least 10 minutes, at least 15 minutes, at least 20 minutes, at least25 minutes or at least 30 minutes.

In yet another aspect, the present disclosure provides a method forpromoting skin rejuvenation, comprising, topically applying abiophotonic composition to a target skin tissue, wherein the biophotoniccomposition comprises at least a first chromophore and a gelling agent;and illuminating said biophotonic composition with light having awavelength that overlaps with an absorption spectrum of the firstchromophore; wherein the gelling agent renders the biophotoniccomposition substantially resistant to leaching such that less than 15%of the total chromophore amount leaches out of the composition intotissue. In certain embodiments, the composition is substantiallyresistant to leaching such that less than 15% of the total chromophoreamount leaches out of the biophotonic composition into tissue during atreatment time in which the composition is topically applied ontotissue. In some embodiments, the biophotonic is substantially resistantto leaching such that less than about 15% of total chromophore contentleaches out of the biophotonic composition during a treatment time of atleast 5 minutes, at least 10 minutes, at least 15 minutes, at least 20minutes, at least 25 minutes or at least 30 minutes.

In yet another aspect, the present disclosure provides a method forcosmetic skin treatment, comprising, topically applying a biophotoniccomposition to a target skin tissue, wherein the biophotonic compositioncomprises at least a first chromophore and gelling agent; andilluminating said biophotonic composition with light having a wavelengththat overlaps with an absorption spectrum of the first chromophore;wherein the gelling agent renders the biophotonic compositionsubstantially resistant to leaching such that less than 15% of the totalchromophore amount leaches out of the composition into tissue. Incertain embodiments, the composition is substantially resistant toleaching such that less than 15% of the total chromophore amount leachesout of the biophotonic composition into tissue during a treatment timein which the composition is topically applied onto tissue. In someembodiments, the biophotonic is substantially resistant to leaching suchthat less than about 15% of total chromophore content leaches out of thebiophotonic composition during a treatment time of at least 5 minutes,at least 10 minutes, at least 15 minutes, at least 20 minutes, at least25 minutes or at least 30 minutes.

In yet another aspect, the present disclosure provides a method fortreatment of periodontal disease, comprising: topically applying abiophotonic composition to a periodontal pocket, wherein the biophotoniccomposition comprises at least a first chromophore and a gelling agent;and illuminating said biophotonic composition with light having awavelength that overlaps with an absorption spectrum of the firstchromophore; wherein the gelling agent renders the biophotoniccomposition substantially resistant to leaching such that less than 15%of the total chromophore amount leaches out of the composition intoperiodontal tissue. In certain embodiments, the composition issubstantially resistant to leaching such that less than 15% of the totalchromophore amount leaches out of the biophotonic composition intoperiodontal tissue during a treatment time in which the composition istopically applied onto periodontal tissue. In some embodiments, thebiophotonic is substantially resistant to leaching such that less thanabout 15% of total chromophore content leaches out of the biophotoniccomposition during a treatment time of at least 5 minutes, at least 10minutes, at least 15 minutes, at least 20 minutes, at least 25 minutesor at least 30 minutes.

In certain embodiments of any method of the present disclosure, thebiophotonic composition is illuminated for any time period per treatmentin which the biophotonic composition is activated, for example 1 to 30minutes. The distance of the light source from the biophotoniccomposition can be any distance which can deliver an appropriate lightpower density to the biophotonic composition and/or the skin tissue, forexample 5, 10, 15 or 20 cm. The biophotonic composition is appliedtopically at any suitable thickness. Typically, the biophotoniccomposition is applied topically to skin or wounds at a thickness of atleast about 2 mm, about 2 mm to about 10 mm.

In certain embodiments, the method of the present disclosure comprises astep of illuminating the biophotonic composition for a period of atleast 30 seconds, 2 minutes, 3 minutes, 5 minutes, 7 minutes, 10minutes, 15 minutes, 20 minutes, 25 minutes, or 30) minutes. In someembodiments, the biophotonic composition is illuminated for a period ofat least 3 minutes.

In certain embodiments of the methods of the present disclosure, thebiophotonic composition is removed from the site of a treatmentfollowing application of light. Accordingly, the biophotonic compositionis removed from the site of treatment within at least 30 seconds, 2minutes, 3 minutes, 5 minutes, 7 minutes, 10 minutes, 15 minutes, 20minutes, 25 minutes or 30 minutes after application. In someembodiments, the biophotonic composition is illuminated for a period ofat least 3 minutes. In some embodiments, the biophotomic composition isremoved after a period of at least 3 minutes post application of thebiophotonic composition to treatment site.

In certain other embodiments, the biophotonic composition is kept inplace for up to one, two or three weeks, and illuminated with lightwhich may include ambient light at various intervals. In this case, thecomposition may be covered up in between exposure to light. For example,the biophotonic composition may be soaked in a dressing and placedinside or over a wound and be left in place for an extended period oftime (e.g. more than one day).

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 depicts absorption of light in the various layers of the skin(Samson et al. Evidence Report Technology Assessment 2004, 111, pages1-97).

FIG. 2 illustrates the Stokes' shift.

FIG. 3 illustrates the absorption and emission spectra of donor andacceptor chromophores. The spectral overlap between the absorptionspectrum of the acceptor chromophore and the emission spectrum of thedonor chromophore is also shown.

FIG. 4 is a schematic of a Jablonski diagram that illustrates thecoupled transitions involved between a donor emission and acceptorabsorbance.

FIG. 5 depicts the experimental setup of an in vitro release test forevaluating leaching of the chromophore(s) of the biophotoniccompositions (Example 6).

FIGS. 6 a and 6 b are absorbance and emission spectra, respectively, ofa composition according to certain embodiments of the present disclosurewhich includes Eosin and Fluorescein in a gel (Example 1).

FIGS. 7 a and 7 b are absorbance and emission spectra, respectively, ofa composition according to certain embodiments of the present disclosurewhich includes Eosin and Fluorescein in an aqueous solution (Example 2).

FIGS. 8 a and 8 b are absorbance and emission spectra, respectively, ofa composition according to certain embodiments of the present disclosurewhich includes Eosin, Fluorescein and Rose Bengal in a gel (Example 3).

FIGS. 9 a and 9 b are absorbance and emission spectra, respectively, ofa composition according to certain embodiments of the present disclosurewhich includes Eosin and Fluorescein in an aqueous solution (Example 4).

FIG. 10 illustrates a summary of inflammatory lesion count and absolutechanges by hemiface (Example 5).

FIG. 11 shows the effect of a biophotonic composition of the disclosureon Ki67 expression (Example 10).

FIG. 12 shows that emitted fluorescence from chromophore in acomposition increases rapidly with increasing composition but slows downto a plateau with further concentration increase for Eosin Y (top) andFluorescein (bottom) (Example 13).

FIG. 13 shows that Eosin and Rose Bengal act in a synergistic manner(Example 14).

FIG. 14 is an emission spectrum showing the intensity over time of thelight being emitted from the biophotonic composition tested in Example5.

FIG. 15 is an emission spectrum showing the intensity over time of thelight being emitted from the biophotonic composition tested in Example7.

DETAILED DESCRIPTION (1) Overview

Phototherapy regimens have been developed to promote wound healing,rejuvenate facial skins and treat various skin disorders. However, thesemethods require direct application of a photosensitive agent (orchromophore) to the target skin and/or uptake of the photosensitiveagent (or chromophore) into the skin cells. As mentioned above, thedirect contact of the photosensitive agent with the tissue can lead toundesired side-effects, including cellular damage/destruction andsystemic or localized toxicity to the patient. Moreover, many existingphototherapy regimens often demonstrate low therapeutic efficacy due to,for example, the poor update of the photosensitive agents into the skincells the target site. For this reason, may regimens require a wait timeof between about one and 72 hours to allow the internalization of thephotosensitizer.

The present disclosure provides biophotonic compositions including aphotoactive exogenous chromophore and methods useful for promoting woundhealing, cosmetic treatment of skin such as skin rejuvenation, treatingacne and treating other skin disorders, treating acute inflammation,which are distinguished from conventional photodynamic therapy.Biophotonic therapy using these compositions does not rely oninternalization of the chromophore into cells or substantial contactwith the cells or target tissues. Therefore, the undesired side effectscaused by direct contact may be reduced, minimized, or prevented. Atmost, the chromophore has surface contact with the tissue to which thecomposition is applied.

(2) Definitions

Before continuing to describe the present disclosure in further detail,it is to be understood that this disclosure is not limited to specificcompositions or process steps, as such may vary. It must be noted that,as used in this specification and the appended claims, the singular form“a”, “an” and “the” include plural referents unless the context clearlydictates otherwise.

As used herein, the term “about” in the context of a given value ofrange refers to a value or range that is within 20%, preferably within10%, and more preferably within 5% of the given value of range.

It is convenient to point out here that “and/or” where used herein is tobe taken as specific disclosure of each of the two specified features orcomponents with or without the other. For example “A and/or B” is to betaken as specific disclosure of each of (i) A, (ii) B and (iii) A and B,just as if each is set out individually herein.

“Biophotonic” means the generation, manipulation, detection andapplication of photons in a biologically relevant context. In otherwords, biophotonic compositions exert their physiological effectsprimarily due to the generation and manipulation of photons.“Biophotonic composition” is a composition as described herein that maybe activated by light to produce photons for biologically relevantapplications.

“Topical composition” means a composition to be applied to bodysurfaces, such as the skin, mucous membranes, vagina, oral cavity,internal surgical wound sites, and the like. A topical composition maybe in the form of, including, but not limited to, a cream, gel,ointment, lotion, levigate, solution, paste, putty, bioadhesive, salve,milk, impregnated material such as a pad, sheet, fabric or fibres,dressings, spray, suspension, foam, or the like.

Terms “chromophore”, “photoactivating agent” and “photoactivator” areused herein interchangeably. A chromophore means a chemical compound,when contacted by light irradiation, is capable of absorbing the light.The chromophore readily undergoes photoexcitation and can then transferits energy to other molecules or emit it as light.

“Photobleaching” means the photochemical destruction of a chromophore.

“Leaching” means the release of one or more components of a biophotoniccomposition (e.g., the chromophore(s) from the composition to thesurrounding environment such as for example the wound site or into thetissue being treated with the composition).

The term “actinic light” is intended to mean light energy emitted from aspecific light source (e.g., lamp, LED, or laser) and capable of beingabsorbed by matter (e.g. the chromophore or photoactivator definedabove). In a preferred embodiment, the actinic light is visible light.

As used herein, a “hygroscopic” substance is a substance capable oftaking up water, for example, by absorption or adsorption even atrelative humidity as low as 50%, at room temperature (e.g. about 25°C.).

“Impermeable membrane” means that the material contained within themembrane is sufficiently or substantially impermeable to the surroundingenvironment such that the migration of such material out of themembrane, and/or the migration of the environmental components (such aswater) into the membrane, is so low as to having substantially noadverse impact on the function or activity of the materials retainedwithin the membrane. The impermeable membrane may be ‘breathable’ inthat gas flow through the membrane is permitted whilst the flow ofliquid is not permitted. The impermeable membrane may also selectivelyallow the migration of some of the materials through the membrane butnot others.

“Wound” means an injury to any tissue, including for example, acute,subacute, delayed or difficult to heal wounds, and chronic wounds.Examples of wounds may include both open and closed wounds. Woundsinclude, for example, burns, incisions, excisions, lesions, lacerations,abrasions, puncture or penetrating wounds, surgical wounds, contusions,hematomas, crushing injuries, ulcers (such as for example pressure,venous, pressure of diabetic), wounds caused by periodontitis(inflammation of the periodontium).

“Skin rejuvenation” means a process of reducing, diminishing, retardingor reversing one or more signs of skin aging. For instance, common signsof skin aging include, but are not limited to, appearance of fine linesor wrinkles, thin and transparent skin, loss of underlying fat (leadingto hollowed cheeks and eye sockets as well as noticeable loss offirmness on the hands and neck), bone loss (such that bones shrink awayfrom the skin due to bone loss, which causes sagging skin), dry skin(which might itch), inability to sweat sufficiently to cool the skin,unwanted facial hair, freckles, age spots, spider veins, rough andleathery skin, fine wrinkles that disappear when stretched, loose skin,or a blotchy complexion. According to the present disclosure, one ormore of the above signs of aging may be reduced, diminished, retarded oreven reversed by the compositions and methods of the present disclosure.

(3) Biophotonic Topical Compositions

The present disclosure provides biophotonic compositions. Biophotoniccompositions are compositions that are, in a broad sense, activated bylight (e.g., photons) of specific wavelength. These compositions containat least one exogenous chromophore which is activated by light andaccelerates the dispersion of light energy, which leads to lightcarrying on a therapeutic effect on its own, and/or to the photochemicalactivation of other agents contained in the composition (e.g.,acceleration in the breakdown process of peroxide (an oxygen-releasingagent) when such compound is present in the composition or at thetreatment site, leading to the formation of oxygen radicals, such assinglet oxygen).

In some aspects, the present disclosure provides biophotoniccompositions comprising at least a first chromophore and a gellingagent, wherein the composition is substantially resistant to leachingsuch that a low chromophore amount leaches out of the biophotoniccomposition into tissue during treatment. In other aspects, the presentdisclosure provides a first composition and a second composition,wherein the first composition comprises an oxygen-releasing agent andthe second composition comprises one or more chromophores, which, whenmixed with the first composition and subsequently activated by light,disperses the light energy, leading to the photochemical activation ofthe oxygen-releasing agent contained in the mixture, which may lead tothe formation of oxygen radicals, such as singlet oxygen.

When a chromophore absorbs a photon of a certain wavelength, it becomesexcited. This is an unstable condition and the molecule tries to returnto the ground state, giving away the excess energy. For somechromophores, it is favorable to emit the excess energy as light whentransforming back to the ground state. This process is calledfluorescence. The peak wavelength of the emitted fluorescence is shiftedtowards longer wavelengths compared to the absorption wavelengths due toloss of energy in the conversion process. This is called the Stokes'shift and is illustrated in FIG. 2 . In the proper environment (e.g., ina biophotonic composition) much of this energy is transferred to theother components of the composition or to the treatment site directly.

Without being bound to theory, it is thought that fluorescent lightemitted by photoactivated chromophores may have therapeutic propertiesdue to its femto-, pico- or nano-second emission properties which may berecognized by biological cells and tissues, leading to favorablebiomodulation. Furthermore, the emitted fluorescent light bas a longerwavelength and hence a deeper penetration into the tissue than theactivating light. Irradiating tissue with such a broad range ofwavelengths, including in some embodiments the activating light whichpasses through the composition, may have different and complementaryeffects on the cells and tissues. Moreover, the generation of oxygenspecies by photoactivated chromophores has been observed by theinventors to cause micro-bubbling within the composition which can havea physical impact on the tissue to which it is applied, for example bydislodging biofilm and debridement of necrotic tissue or providing apressure stimulation. The biofilm can also be pre-treated with anoxygen-releasing agent to weaken the biofilm before treating with thecomposition of the present disclosure.

The biophotonic compositions of the present disclosure are substantiallytransparent/translucent and/or have high light transmittance in order topermit light dissipation into and through the composition. In this way,the area of tissue under the composition can be treated both with thefluorescent light emitted by the composition and the light irradiatingthe composition to activate it. The % transmittance of the biophotoniccomposition can be measured in the range of wavelengths from 250 nm to800 nm using, for example, a Perkin-Elmer Lambda 9500 series UV-visiblespectrophotometer. In some embodiments, transmittance of thecompositions disclosed herein is measured at 460 nm.

As transmittance is dependent upon thickness, the thickness of eachsample can be measured with calipers prior to loading in thespectrophotometer. Transmittance values can be normalized to a thicknessof 100 μm (or any thickness) according to:

${{F_{T - {corr}}\left( {\lambda,t_{2}} \right)} = {\left\lbrack {{e^{- \sigma_{t}}(\lambda)}t_{1}} \right\rbrack^{\frac{t_{2}}{t_{1}}} = \left\lbrack {F_{T - {corr}}\left( {\lambda,t_{1}} \right)} \right\rbrack^{\frac{t_{2}}{t_{1}}}}},$

where t₁=actual specimen thickness, t₂=thickness to which transmittancemeasurements can be normalized.

In some embodiments, the biophotonic composition has a transparency ortranslucency that exceeds 15%, 20%, 30%, 40%, 50%, 55%, 60%, 65%, 70%,75%, 80%, or 85% at 460 nm. In some embodiments, the transparencyexceeds 70% at 460 nm, 86% at 460 nm, 87% at 460 nm, 88% at 460 nm, 89%at 460 nm, 90% at 460 nm, 91% at 460 nm, 92% at 460 nm, 93% at 460 nm,94% at 460 nm, 95% at 460 nm, 96% at 460 nm, 97% at 460 nm, 98% at 460nm or 99% at 460 nm.

The biophotonic compositions of the present disclosure are for topicaluses. These compositions may be described based on the components makingup the composition. Additionally or alternatively, the compositions ofthe present disclosure have functional and structural properties andthese properties may also be used to define and describe thecompositions. Individual components of the composition of the presentdisclosure are detailed as below.

(a) Chromophores

The biophotonic topical compositions of the present disclosure compriseone or more chromophores, which can be considered exogenous, e.g., arenot naturally present in skin or tissue.

Suitable chromophores can be fluorescent dyes (or stains), althoughother dye groups or dyes (biological and histological dyes, foodcolorings, carotenoids, naturally occurring fluorescent and other dyes)can also be used. Suitable photoactivators can be those that areGenerally Regarded As Safe (GRAS). Photoactivators which are not welltolerated by the skin or other tissues can be included in thebiophotonic composition in an encapsulated form.

In certain embodiments, the biophotonic topical composition of thepresent disclosure comprises a first chromophore which undergoes partialor complete photobleaching upon application of light. By photobleachingis meant a photochemical destruction of the chromophore which cangenerally be visualized as a loss of color.

In some embodiments, the first chromophore absorbs at a wavelength inthe range of the visible spectrum, such as at a wavelength of about380-800 nm, 380-700, or 380-600 nm. In other embodiments, the firstchromophore absorbs at a wavelength of about 200-800 nm, 200-700 nm,200-600 nm or 200-500 nm. In one embodiment, the first chromophoreabsorbs at a wavelength of about 200-600 nm. In some embodiments, thefirst chromophore absorbs light at a wavelength of about 200-300 nm,250-350 nm, 300-400 nm, 350-450 nm, 400-500 nm, 400-600 nm, 450-650 nm,600-700 nm, 650-750 nm or 700-800 nm.

It will be appreciated to those skilled in the art that opticalproperties of a particular chromophore may vary depending on thechromophore's surrounding medium. Therefore, as used herein, aparticular chromophore's absorption and/or emission wavelength (orspectrum) corresponds to the wavelengths (or spectrum) measured in abiophotonic composition of the present disclosure.

The biophotonic compositions disclosed herein may include at least oneadditional chromophore. Combining chromophores may increasephoto-absorption by the combined dye molecules and enhance absorptionand photo-biomodulation selectivity. This creates multiple possibilitiesof generating new photosensitive, and/or selective chromophoresmixtures.

When such multi-chromophore compositions are illuminated with light,energy transfer can occur between the chromophores. This process, knownas resonance energy transfer, is a photophysical process through whichan excited ‘donor’ chromophore (also referred to herein as firstchromophore) transfers its excitation energy to an ‘acceptor’chromophore (also referred to herein as second chromophore). Theefficiency and directedness of resonance energy transfer depends on thespectral features of donor and acceptor chromophores. In particular, theflow of energy between chromophores is dependent on a spectral overlapreflecting the relative positioning and shapes of the absorption andemission spectra. For energy transfer to occur the emission spectrum ofthe donor chromophore overlap with the absorption spectrum of theacceptor chromophore (FIG. 3 ).

Energy transfer manifests itself through decrease or quenching of thedonor emission and a reduction of excited state lifetime accompaniedalso by an increase in acceptor emission intensity. FIG. 4 is aJablonski diagram that illustrates the coupled transitions involvedbetween a donor emission and acceptor absorbance.

To enhance the energy transfer efficiency, the donor chromophore shouldhave good abilities to absorb photons and emit photons. Furthermore, itis thought that the more overlap there is between the donorchromospheres' emission spectra and the acceptor chromophore'sabsorption spectra, the better a donor chromophore can transfer energyto the acceptor chromophore.

In certain embodiments, the biophotonic topical composition of thepresent disclosure further comprises a second chromophore. In someembodiments, the first chromophore has an emission spectrum thatoverlaps at least about 80%, 50%, 40%, 30%, 20%, 10% with an absorptionspectrum of the second chromophore. In one embodiment, the firstchromophore has an emission spectrum that overlaps at least about 20%with an absorption spectrum of the second chromophore. In someembodiments, the first chromophore has an emission spectrum thatoverlaps at least 1-10%, 5-15%, 10-20%, 15-25%, 20-30%, 25-35%, 30-40%,35-45%, 50-60%, 55-65% or 60-70% with an absorption spectrum of thesecond chromophore.

% spectral overlap, as used herein, means the % overlap of a donorchromophore's emission wavelength range with an acceptor chromophore'sabsorption wavelength rage, measured at spectral full width quartermaximum (FWQM). For example, FIG. 3 shows the normalized absorption andemission spectra of donor and acceptor chromophores. The spectral FWQMof the acceptor chromophore's absorption spectrum is from about 60 nm(515 nm to about 575 nm). The overlap of the donor chromophore'sspectrum with the absorption spectrum of the acceptor chromophore isabout 40 nm (from 515 nm to about 555 nm). Thus, the % overlap can becalculated as 40 nm/60 nm×100=66.6%.

In some embodiments, the second chromophore absorbs at a wavelength inthe range of the visible spectrum. In certain embodiments, the secondchromophore has an absorption wavelength that is relatively longer thanthat of the first chromophore within the range of about 50-250, 25-150or 10-100 nm.

As discussed above, the application of light to the compositions of thepresent disclosure can result in a cascade of energy transfer betweenthe chromophores. In certain embodiments, such a cascade of energytransfer provides photons that penetrate the epidermis, dermis and/ormucosa at the target tissue, including, such as, a site of wound, or atissue afflicted with acne or a skin disorder. In some embodiments, sucha cascade of energy transfer is not accompanied by concomitantgeneration of heat. In some other embodiments. the cascade of energytransfer does not result in tissue damage.

Optionally, when the biophotonic topical composition comprises a firstand a second chromophore, the first chromophore is present in an amountof about 0.01-40% per weight of the composition, and the secondchromophore is present in an amount of about 0.001-40% per weight of thecomposition. In certain embodiments, the total weight per weight ofchromophore or combination of chromophores may be in the amount of about0.01-40.001% per weight of the composition. In certain embodiments, thefirst chromophore is present in an amount of about 0.01-1%, 0.01-2%,0.05-1%, 0.05-2%, 1-5%, 2.5-7.5%, 5-10%, 7.5-12.5%, 10-15%, 12.5-17.5%,15-20%, 17.5-22.5%, 20-25%, 22.5-27.5%, 25-30%, 27.5-32.5%, 30-35%,32.5-37.5%, or 35-40% per weight of the composition. In certainembodiments, the second chromophore is present in an amount of about0.001-1%, 0.001-2%, 0.001-0.01%, 0 01-0.1%, 0.1-1.0%, 1-2%, 1-5%,2.5-7.5%, 5-10%, 7.5-12.5%, 10-15%, 12.5-17.5%, 15-20%, 17.5-22.5%,20-25%, 22.5-27.5%, 25-30%, 27.5-32.5%, 30-35%, 32.5-37.5%, or 35-40%per weight of the composition. In certain embodiments, the total weightper weight of chromophore or combination of chromophores may be in theamount of about 0.01-1%, 0.01-2%, 0 05-2%, 0.5-1%, 0.5-2%, 1-5%,2.5-7.5%, 5-10%, 7.5-12.5%, 10-15%, 12.5-17.5%, 15-20%, 17.5-22.5%,20-25%, 22.5-27.5%, 25-30%, 27.5-32.5%, 30-35%, 32.5-37.5%, or 35-40.05% per weight of the composition.

In some embodiments, the chromophore or chromophores are selected suchthat their emitted fluorescent light, on photoactivation, is within oneor more of the green, yellow, orange, red and infrared portions of theelectromagnetic spectrum, for example having a peak wavelength withinthe range of about 490 am to about 800 nm. In certain embodiments, theemitted fluorescent light has a power density of between 0.005 to about10 mW/cm², about 0.5 to about 5 mW/cm².

Suitable chromophores that may be used in the biophotonic topicalcompositions of the present disclosure include, but are not limited tothe following:

Chlorophyll Dyes

Exemplary chlorophyll dyes include but are not limited to chlorophyll a;chlorophyll b; oil soluble chlorophyll bacteriochlorophyll a;bacteriochlorophyll b; bacteriochlorophyll c; bacteriochlorophyll d;protochlorophyll; protochlorophyll a; amphiphilic chlorophyll derivative1; and amphiphilic chlorophyll derivative 2.

Xanthene Derivatives

Exemplary xanthene dyes include but are not limited to Eosin B(4,5-dibromo,2′,7′-dinitr-o-fluorescein, dianion); eosin Y; cosin Y(2′,4′,5′,7′-tetrabromo-fluorescein, dianion); eosin(2′,4′,5′,7′-tetrabromo-fluorescein, dianion); eosin(2′,4′,5′,7′-tetrabromo-fluorescein, dianion) methyl ester; cosin(2′,4′,5′,7-tetrabromo-fluorescein, monoamion) p-isopropylbenzyl ester;eosin derivative (2′,7′-dibromo-fluorescein, dianion); cosin derivative(4′,5′-dibromo-fluorescein, dianion); eosin derivative(2′,7′-dichloro-fluorescein, dianion); eosin derivative(4′,5′-dichloro-fluorescein, dianion); cosin derivative(2′,7′-diiodo-fluorescein, dianion); eosin derivative(4′,5′-diiodo-fluorescein, dianion); eosin derivative(tribromo-fluorescein, dianion): eosin derivative(2′,4′,5′,7′-tetrachlor-o-fluorescein, dianion); cosin; cosindicetylpyridinium chloride ion pair, erythrosin B(2′,4′,5′,7′-tetraiodo-fluorescein, dianion); erythrosin; erythrosindianion; erythrosin B, fluorescein; fluorescein dianion; phloxin B(2′,4′,5′,7′-tetrabromo-3,4,5,6-tetrachloro-fluorescein, dianion);phloxin B (tetrachloro-tetrabromo-fluorescein); phloxine B; rose bengal(3,4,5,6-tetrachloro-2′,4′,5′,7′-tetraiodofluorescein, dianion); pyroninG, pyronin J, pyronin Y; Rhodamine dyes such as rhodamines include4,5-dibromo-rhodamine methyl ester; 4,5-dibromo-rhodamine n-butyl ester,rhodamine 101 methyl ester; rhodamine 123; rhodamine 6G; chodamine 6Ghexyl ester; tetrabromo-rhodamine 123, and tetramethyl-rhodamine ethylester.

Methylene Blue Dyes

Exemplary methylene blue derivatives include but are not limited to1-methyl methylene blue; 1,9-dimethyl methylene blue; methylene blue;methylene blue (16 mu.M); methylene blue (14 .mu.M); methylene violet;bromomethylene violet; 4-iodomethylene violet,1,9-dimethyl-3-dimethyl-amino-7-diethyl-a-mino-phenothiazine, and1.9-dimethyl-3-diethylamino-7-dibutyl-amino-phenol-biazine.

Azo Dyes

Exemplary azo (or diazo-) dyes include but are not limited to methylviolet, neutral red, para red (pigment red 1), amaranth (Azorubine S),Carmoisine (azorubine, food red 3, acid red 14), allura red AC (FD&C40), tartrazine (FD&C Yellow 5), orange G (acid orange 10), Ponceau 4R(food red 7), methyl red (acid red 2), and murexide-ammonium purpurate.

In some aspects of the disclosure, the one or more chromophores of thebiophotonic composition disclosed herein can be independently selectedfrom any of Acid black 1, Acid blue 22, Acid blue 93, Acid fuchsin, Acidgreen, Acid green 1, Acid green 5, Acid magenta, Acid orange 10, Acidred 26, Acid red 29, Acid red 44, Acid red 51, Acid red 66, Acid red 87,Acid red 91, Acid red 92, Acid red 94, Acid red 101, Acid red 103, Acidroseine, Acid rubin, Acid violet 19, Acid yellow 1, Acid yellow 9, Acidyellow 23, Acid yellow 24, Acid yellow 36, Acid yellow 73, Acid yellowS, Acridine orange, Acriflavine, Alcian blue, Alcian yellow, Alcoholsoluble eosin, Alizarin, Alizarin blue 2RC, Alzarin carmine, Alizarincyanin BBS, Alizarol cyanin R, Alizarin red S, Alizarin purpurin,Alununon, Amido black 10B, Amidoschwarz, Aniline blue WS, Anthraceneblue SWR, Auramine O, Azocannine B, Azocarmine O, Azoic diazo S, Azoicdiazo 48, Azure A, Azure B, Azure C, Basic blue 8, Basic blue 9, Basicblue 12, Basic blue 15, Basic blue 17, Basic blue 20, Basic blue 26,Basic brown 1, Basic fuchsin, Basic green 4, Basic orange 14, Basic red2 (Saffranin O), Basic red 5, Basic red 9, Basic violet 2, Basic violet3, Basic violet 4, Basic violet 10, Basic violet 14, Basic yellow 1,Basic yellow 2, Biebrich scarlet, Bismarck brown Y, Brilliant crystalscarlet 6R, Calcium red, Carmine, Carminic acid (acid red 4), Celestineblue B, China blue, Cochmeal, Coelestine blue, Chrome violet CG,Chromotrope 2R, Chromoxane cyanin R, Congo corinth, Congo red, Cottonblue, Cotton red, Croceine scarlet, Crocin, Crystal ponceau 6R, Crystalviolet, Dahlia, Diamond green B, DiOC6, Direct blue 14, Direct blue 58,Direct red, Direct red 10, Direct red 28, Direct red 80, Direct yellow7, Eosin B, Eosin Bluish, Eosin, Eosin Y, Eosin yellowish, Eosinol, Eriegarnet B, Eriochrome cyanin R, Erythrosin B, Ethyl eosin, Ethyl green,Ethyl violet, Evans blue, Fast blue B, Fast green FCF, Fast red B, Fastyellow, Fluorescein, Food green 3, Callein, Gallamine blue, Gallocyanin,Gentian violet, Haematein, Haematine, Haematoxylin, Helio fast robinBBL, Helvetia blue, Hematein, Hematine, Hematoxylin, Hoffman's violet,Imperial red, Indocyanin green, Ingrain blue, Ingrain blue 1, Ingrainyellow 1, INT, Kermes, Kermesic acid, Kernechtrot, Lac, Laccaic acid,Lauth's violet, Light green, Lissamine green SF, Luxol fast blue,Magenta 0, Magenta 1, Magenta II, Magenta III, Malachite green,Manchester brown, Martius yellow, Merbromin, Mercurochrome, Metanilyellow, Methylene azure A, Methylene azure B, Methylene azure C,Methylene blue, Methyl blue, Methyl green, Methyl violet, Methyl violet2B, Methyl violet 10B, Mordant blue 3, Mordant blue 10, Mordant blue 14,Mordant blue 23, Mordant blue 32, Mordant blue 45, Mordant red 3,Mordant red 11, Mordant violet 25, Mordant violet 39 Naphthol blueblack, Naphthol green B, Naphthol yellow S, Natural black 1, Naturalred, Natural red 3, Natural red 4, Natural red 8, Natural red 16,Natural red 25, Natural red 28. Natural yellow 6, NBT, Neutral red, Newfuchsin, Niagara blue 3B, Night blue, Nile blue, Nile blue A, Nile blueoxazone, Nile blue sulphate, Nile red, Nitro BT, Nitro blue tetrazolium,Nuclear fast red, Oil red O, Orange G, Orcein, Pararosanilin, PhloxineB, phycobilins, Phycocyanins, Phycoerythrins, Phycoerythrincyanin (PEC),Phthalocyanmes, Picric acid, Ponceau 2R, Ponceau 6R, Ponceau B, Ponceaude Xylidine, Ponceau S, Primula, Purpurin, Pyronin B, Pyronin O, PyroninY, Rhodamine B, Rosanilin, Rose bengal, Saffron, Safranin O, Scarlet R,Scarlet red, Scharlach R, Shellac, Sirius red F3B, Solochrome cyanin R,Soluble blue, Solvent black 3, Solvent blue 38, Solvent red 23, Solventred 24, Solvent red 27, Solvent red 45, Solvent yellow 94, Spiritsoluble eosin, Sudan III, Sudan IV, Sudan black B, Sulfur yellow S,Swiss blue, Tartrazine, Thioflavine S, Thioflavine T, Thionin, Toluidineblue. Toluyline red, Tropaeoli C, Trypaflavine, Trypan blue, Uranin,Victoria blue 4R, Victoria blue B, Victoria green B, Water blue 1, Watersoluble eosin, Xylidine ponceau, or Yellowish eosin.

In certain embodiments, the composition of the present disclosureincludes any of the chromophores listed above, or a combination thereof,so as to provide a biophotonic impact at the application site. This is adistinct application of these agents and differs from the use ofchromophores as simple stains or as a catalyst for photo-polymerization.

In some embodiments, the composition includes Eosin Y as a firstchromophore and any one or more of Rose Bengal, Erythrosin, Phloxine Bas a second chromophore. It is believed that these combinations have asynergistic effect as Eosin Y can transfer energy to Rose Bengal,Erythrosin or Phloxine B when activated. This transferred energy is thenemitted as fluoresence or by production of reactive oxygen species. Thisabsorbed and re-emitted light is thought to be transmitted throughoutthe composition, and also to be transmitted into the site of treatment.

In further embodiments, the composition includes the followingsynergistic combinations: Eosin Y and Fluorescein; Fluorescein and RoseBengal; Erythrosine in combination with Eosin Y, Rose Bengal orFluorescein; Phloxine B in combination with one or more of Eosin Y, RoseBengal, Fluorescein and Erythrosine. Other synergistic chromophorecombinations are also possible.

By means of synergistic effects of the chromophore combinations in thecomposition, chromophores which cannot normally be activated by anactivating light (such as a blue light from an LED) can be activatedthrough energy transfer from chromophores which are activated by theactivating light. In this way, the different properties ofphotoactivated chromophores can be harnessed and tailored according tothe cosmetic or the medical therapy required.

For example, Rose Bengal can generate a high yield of singlet oxygenwhen photoactivated in the presence of molecular oxygen, however it hasa low quantum yield m terms of emitted fluorescent light. Rose Bengalhas a peak absorption around 540 nm and so is normally activated bygreen light. Eosin Y has a high quantum yield and can be activated byblue light. By combining Rose Bengal with Eosin Y, one obtains acomposition winch can emit therapeutic fluorescent light and generatesinglet oxygen when activated by blue light. In this case, the bluelight photoactivates Eosin Y which transfers some of its energy to RoseBengal as well as emitting some energy as fluorescence.

(b) Gelling Agent

The present disclosure provides biophotonic topical compositions thatcomprise at least a first chromophore and a gelling agent, wherein thegelling agent provides a barrier such that the chromophore(s) of thebiophotonic topical compositions are substantially not in contact withthe target tissue.

As used herein, “leaching” means the release of one or more componentsof a biophotonic composition (e.g., the chromophore(s)) from thecomposition to the surrounding environment such as for example the woundsite or into the tissue being treated with the composition). Therefore,the gelling agent present in the biophotonic compositions of the presentdisclosure renders the compositions substantially resistant to leachingsuch that the chromophore(s) or photosensitive agent(s) of thebiophotonic topical compositions are not in substantial contact with thetarget tissue.

In certain embodiments, the biophotonic topical composition allows lessthan 30%, 25%, 20%, 15%, 10%, 5%, 1%, 0.8%, 0.5% or 0.1%, or essentiallynone of said chromophore content to leach out of the biophotoniccomposition.

In some embodiments, the biophotonic composition limits leaching of thefirst chromophore such that less than 15% of total chromophore amountcan leach out into tissue during a treatment time in which thecomposition is topically applied onto tissue and illuminated with light.In some embodiments, the biophotomic composition limits leaching of thefirst chromophore such that less than 30%, 25%, 20%, 15%, 10%, 5%, 1%,0.8%, 0.5% or 0.1% or essentially 0% of total chromophore amount canleach out into tissue during a treatment time in which the compositionis topically applied onto tissue and illuminated with light. In someembodiments, the treatment time is at least about 5 minutes, at leastabout 10 minutes, at least about 15 minutes, at least about 20 minutes,at least about 25 minutes or at least about 30 minutes.

The extent of chromophore leaching out of the biophotonic compositionand into the surrounding environment may be assessed using variousmethods known in the art, including but not limited to, the testsdescribed in the Examples.

In some embodiments, leaching is determined by placing the biophotoniccomposition in contact with an aqueous solution through a porousmembrane for a period of time corresponding to a desired treatment time.The extent of chromophore leaching can then be assessed visually, forexample, by noting a color change of the aqueous solution, orquantitatively, for example, by using a spectrophotometer to measure theabsorption of the solution. In some embodiments, a biophotoniccomposition of the present disclosure allows less than 30%, 25%, 20%,15%, 10%, 5%, 1%, 0.8%, 0.5% or 0.1% or essentially 0% of the totalchromophore amount to leach out of the biophotonic composition asthrough a porous membrane into an aqueous solution when the biophotoniccomposition is placed in contact with the aqueous solution through theporous membrane for a time corresponding to a desired treatment time. Incertain embodiments, the time corresponding to a treatment time is atleast about 5 minutes, at least about 10 minutes, 15 minutes, 20minutes, 25 minutes or 30 minutes.

In some embodiments, staining is determined by visually assessingwhether the biophotonic composition colorizes white test paper saturatedwith 70% by volume ethanol/30% by volume water solution placed incontact with the biophotonic composition for a period of timecorresponding to a desired treatment time. In some embodiments, abiophotonic composition of the present disclosure does not visuallycolorize white test paper saturated with a 70% by volume ethanol/30% byvolume water solution placed in contact with the biophotonic compositionunder atmospheric pressure for a time corresponding to a desiredtreatment. time In certain embodiments, the time corresponding to atreatment time is at least about 5 minutes, at least about 10 minutes,15 minutes, 20 minutes, 25 minutes or 30 minutes.

A gelling agent for use according to the present disclosure may compriseany ingredient suitable for use in a topical biophotonic formulation asdescribed herein. The gelling agent according to various embodiments ofthe present disclosure may include, but not limited to, polyalkyleneoxides, particularly polyethylene glycol and poly(ethyleneoxide)-poly(propylene oxide) copolymers, including block and randomcopolymers; polyols such as glycerol, polyglycerol (particularly highlybranched polyglycerol), propylene glycol and trimethylene glycolsubstituted with one or more polyalkylene oxides, e.g., mono-, di- andtri-polyoxyethylated glycerol, mono- and di-polyoxy-ethylated propyleneglycol, and mono- and di-polyoxyethylated trimethylene glycol,polyoxyethylated sorbitol, polyoxyethylated glucose, acrylic acidpolymers and analogs and copolymers thereof, such as polyacrylic acidper se, polymethacrylic acid, poly(hydroxyethylmethacrylate), poly(hydroxyethylacrylate), poly(methylalkylsulfoxide methacrylate),poly(methylalkylsulfoxide acrylate) and copolymers of any of theforegoing, and/or with additional acrylate species such as antinoethylacrylate and mono-2-(acryloxy)-ethyl succinate, polymaleic acid;poly(acrylamides) such as polyacrylamide per se, poly(methacrylamide),poly(dimethylacrylamide), and poly(N-isopropyl-acrylamide);poly(olefinic alcohol)s such as poly(vinyl alcohol); poly(N-vinyllactams) such as poly(vinyl pyrrolidone), poly(N-vinyl caprolactam), andcopolymers thereof, polyoxazolines, including poly(methyloxazoline) andpoly(ethyloxazoline); and polyvinylamines.

The gelling agent according to certain embodiments of the presentdisclosure may include a polymer selected from any of synthetic orsemi-synthetic polymeric materials, polyacrylate copolymers, cellulosederivatives and polymethyl vinyl ether/maleic anhydride copolymers. Insome embodiments, the hydrophilic polymer comprises a polymer that is ahigh molecular weight (i.e., molar masses of more than about 5,000, andin some instances, more than about 10,000, or 100,000, or 1,000,000)and/or cross-linked polyacrylic acid polymer. In some embodiments, thepolymer is a polyacrylic acid polymer and has a viscosity in the rangeof about 15,000-100,000, 15,000-90,000, 15,000-80,000, 20,000-80,000,20,000-70,000, 20,000-40,000 cP. In certain embodiment, the polymer is ahigh molecular weight, and/or cross-linked polyacrylic acid polymer,where the polyacrylic acid polymer bas a viscosity in the range of about15,000-80,000 cP.

In some embodiments, the gelling agent comprises a carbomer. Carbomersare synthetic high molecular weight polymer of acrylic acid that arecrosslinked with either allylsucrose or allylethers of pentaerythritolhaving a molecular weight of about 3×10⁶. The gelation mechanism dependson neutralization of the carboxylic acid moiety to form a soluble salt.The polymer is hydrophilic and produces sparkling clear gels whenneutralized. Carbomer gels possess good thermal stability in that gelviscosity and yield value are essentially unaffected by temperature. Asa topical product, carbomer gels possess optimum rheological properties.The inherent pseudoplastic flow permits immediate recovery of viscositywhen shear is terminated and the high yield value and quick break makeit ideal for dispensing. Aqueous solution of Carbopol® is acidic innature due to the presence of free carboxylic acid residues.Neutralization of this solution cross-links and gelatinizes the polymerto form a viscous integral structure of desired viscosity.

Carbomers are available as fine white powders which disperse in water toform acidic colloidal suspensions (a 1% dispersion has approx. pH 3) oflow viscosity. Neutralization of these suspensions using a base, forexample sodium, potassium or ammonium hydroxides, low molecular weightamines and alkanolamines, results in the formation of translucent gels.Nicotine salts such as nicotine chloride form stable water-solublecomplexes with carbomers at about pH 3.5 and are stabilized at anoptimal pH of about 5.6.

In some embodiments of the disclosure, the carbomer is Carbopol. Suchpolymers are commercially available from B.F. Goodrich or Lubrizol underthe designation Carbopol® 71G NF, 420, 430, 475, 488, 493, 910, 934,934P, 940, 971PNF, 974P NF, 980 NF, 981 NF and the like. Carbopols areversatile controlled-release polymers, as described by Brock(Pharmacotherapy, 14:430-7 (1994)) and Durrani (Pharmaceutical Res.(Supp.) 8:S˜135 (1991)), and belong to a family of carbomers which aresynthetic, high molecular weight, non-linear polymers of acrylic acid,crosslinked with polyalkenyl polyether. In some embodiments, thecarbomer is Carbopol® 974P NF, 980 NF, 5984 EP, ETD 2020NF, Ultrez 10NF, 934 NF, 934P NF or 940 NF. In certain embodiments, the carbomer isCarbopol® 980 NF, ETD 2020 NF, Ultrez 10 NF, Ultrez 21 or 1382 Polymer,1342 NF, 940 NF.

In certain embodiments, the gelling agent comprises a hygroscopicmaterial. The hygroscopic material may include, but is not limited to,glucosamine, glycosaminoglycan, poly(vinyl alcohol),poly(2-hydroxyethylmethylacrylate), polyethylene oxide, collagen,chitosan, alginate, a poly(acrylonitrile)-based hydrogel, poly(ethyleneglycol)/poly(acrylic acid) interpenetrating polymer network hydrogel,polyethylene oxide-poly butylene terephthalate, hyaluronic acid,high-molecular-weight polyacrylic acid, poly(hydroxy ethylmethacrylate),polyethylene glycol), tetraethylene glycol diacrylate, polyethyleneglycol methacrylate, and poly(methyl acrylate-co-hydroxyethyl acrylate).

The one or more gelling agents can be selected according to theirability to prevent leaching. For example, gelling agents which canincrease the viscosity of the biophotonic composition can be selected.In some embodiments, the viscosity of the biophotonic composition is15,000-100,000, 15,000-90,000, 15,000-80,000, 20,000-80,000,20,000-70,000, 20,000-40,000 cP. A composition with sufficiently highviscosity parameters can prevent or limit the leaching of chromophoresfrom the composition. Gelling agents which include lipids or othercoating agents which can coat the chromophores can also be used to limitor prevent leaching. Alternatively or in addition to the aforesaid,gelling agents which are hygroscopic and/or hydrophilic may be used fortheir water attracting properties, which may also prevent or limitleaching of the chromophore.

Viscosity of the biophotonic compositions of the present disclosure maybe measured using a cone/plate viscometer (Wells-Brookfield). A CP-51cone may be used and viscosity is measured at a speed of 2 rpm andmaking sure that the torque is >10%. Spindle must rotate at least 5times before a viscosity reading is taken.

The biophotonic composition of the present disclosure may be furtherencapsulated, e.g. in a membrane. Such a membrane may be transparent,and/or substantially, or fully impermeable. The membrane may beimpermeable to liquid but permeable to gases such as air. In certainembodiments, the composition may form a membrane that encapsulates thechromophore(s) of the biophotonic topical composition, where themembrane may be substantially impermeable to liquid and/or gas. Incertain embodiments, the biophotonic composition is a liquidencapsulated by a membrane, wherein the membrane is sufficientlyresistant to chromophore leaching such that less than 15% of the totalchromophore amount leaches out of the encapsulated composition. Themembrane may be formed of one or more lipidic agents.

(c) Oxygen-Releasing Agents

According to certain embodiments, the compositions of the presentdisclosure may optionally further comprise one or more additionalcomponents, such as oxygen-releasing agents. For instance, thebiophotonic topical composition of the present disclosure may optionallycomprise oxygen-releasing agents as a source of oxygen. Peroxidecompounds are oxygen-releasing agents that contain the peroxy group(R—O—O—R), which is a chainlike structure containing two oxygen atoms,each of which is bonded to the other and a radical or some element.

When a biophotonic composition of the present disclosure comprising anoxygen-releasing agent is illuminated with light, the chromophore(s) areexcited to a higher energy state. When the chromophore(s)' electronsreturn to a lower energy state, they emit photons with a lower energylevel, thus causing the emission of light of a longer wavelength(Stokes' shift). In the proper environment, some of this energy releaseis transferred to oxygen or the reactive hydrogen peroxide and causesthe formation of oxygen radicals, such as singlet oxygen. The singletoxygen and other reactive oxygen species generated by the activation ofthe biophotonic composition are thought to operate in a hormeticfashion. That is, a health beneficial effect that is brought about bythe low exposure to a normally toxic stimuli (e.g. reactive oxygen), bystimulating and modulating stress response pathways in cells of thetargeted tissues. Endogenous response to exogenous generated freeradicals (reactive oxygen species) is modulated in increased defensecapacity against the exogenous free radicals and induces acceleration ofhealing and regenerative processes. Furthermore, activation of thecomposition can also produce an antibacterial effect. The extremesensitivity of bacteria to exposure to free radicals makes thecomposition of the present disclosure a de facto bactericidalcomposition.

As stated above, the generation of oxygen species by the composition insome embodiments is accompanied by the micro-bubbling which cancontribute to debridement or dislodging of biofilm at the site ofapplication. This can allow for the improved penetration of theactivating and/or fluorescence light to the treatment site for exampleto deactivate bacterial colonies leading to their reduction in number.

Suitable oxygen-releasing agents that may be included in the compositioninclude, but are not limited to.

Hydrogen peroxide (H₂O₂) is the starting material to prepare organicperoxides. H₂O₂ is a powerful oxygen-releasing agent, and the uniqueproperty of hydrogen peroxide is that it breaks down into water andoxygen and does not form any persistent, toxic residual compound.Hydrogen peroxide for use in this composition can be used in a gel, forexample with 6% hydrogen peroxide. A suitable range of concentrationover which hydrogen peroxide can be used in the present composition isfrom about 0.1% to about 6%.

Urea hydrogen peroxide (also known as urea peroxide, carbamide peroxideor percarbamide) is soluble in water and contains approximately 35%hydrogen peroxide. Carbamide peroxide for use in this composition can beused as a gel, for example with 16% carbamide peroxide that represents5.6% hydrogen peroxide, or 12% carbamide peroxide. A suitable range ofconcentration over which urea peroxide can be used in the presentcomposition is from about 0.3% to about 16%. Urea peroxide breaks downto urea and hydrogen peroxide in a slow-release fashion that can beaccelerated with heat of photochemical reactions. The released urea[carbamide, (NH₂)CO₂)], is highly soluble in water and is a powerfulprotein denaturant. It increases solubility of some proteins andenhances rehydration of the skin and/or mucosa.

Benzoyl peroxide consists of two benzoyl groups (benzoic acid with the Hof the carboxylic acid removed) joined by a peroxide group. It is foundin treatments for acne, in concentrations varying from 2.5% to 10%. Thereleased peroxide groups are effective at killing bacteria. Benzoylperoxide also promotes skin turnover and clearing of pores, whichfurther contributes to decreasing bacterial counts and reduce acneBenzoyl peroxide breaks down to benzoic acid and oxygen upon contactwith skin, neither of which is toxic. A suitable range of concentrationover which benzoyl peroxide can be used in the present composition isfrom about 2.5% to about 5%.

Specific oxygen-releasing agents that that are preferably used in thematerials or methods of this disclosure include, but are not limited tohydrogen peroxide, carbamide peroxide. or benzoyl peroxide. Inclusion ofother forms of peroxides (e.g. organic or inorganic peroxides) should beavoided due to their increased toxicity and their unpredictable reactionwith the photodynamic energy transfer. Oxygen-releasing agents can beprovided in powder, liquid or gel form. Alternatively, theoxygen-releasing agents may also be applied to the tissue siteseparately to the composition. Alternatively, the composition mayinclude an amount of oxygen-releasing agent, which is augmented by theseparate application of oxygen-releasing agents to the treatment site.

In the compositions and methods of the present disclosure, additionalcomponents may optionally be included, or used in combination with thebiophotonic compositions as described herein. Such additional componentsinclude, but are not limited to, healing factors, growth factors,antimicrobials, wrinkle fillers (e.g. botox, hyaluronic acid orpolylactic acid), collagens, anti-virals, anti-fungals, antibiotics,drugs, and/or agents that promote collagen synthesis. These additionalcomponents may be applied to the wound, skin or mucosa in a topicalfashion, prior to, at the same time of, and/or after topical applicationof the biophotonic composition of the present disclosure, and may alsobe systemically administered. Suitable healing factors, antimicrobials,collagens, and/or agents that promote collagen synthesis are discussedbelow:

(d) Healing Factors

Healing factors comprise compounds that promote or enhance the healingof regenerative process of the tissues on the application site of thecomposition. During the photoactivation of the composition of thepresent disclosure, there is an increase of the absorption of moleculesat the treatment site by the skin, wound or the mucosa. An augmentationin the blood flow at the site of treatment is observed for an extentperiod of time. An increase in the lymphatic drainage and a possiblechange in the osmotic equilibrium due to the dynamic interaction of thefree radical cascades can be enhanced or even fortified with theinclusion of healing factors. Suitable healing factors include, but arenot limited to.

Hyaluronic acid (Hyaluronan, hyaluronate): is a non-sulfatedglycosaminoglycan, distributed widely throughout connective, epithelialand neural tissues. It is one of the primary components of theextracellular matrix, and contributes significantly to cellproliferation and migration. Hyaluronan is a major component of theskin, where it is involved in tissue repair. While it is abundant inextracellular matrices, it contributes to tissues hydrodynamics,movement and proliferation of cells and participates in a wide number ofcell surface receptor interactions, notably those including primaryreceptor CD44. The hyaluronidases enzymes degrade hyaluronan. There areat least seven types of hyaluronidase-like enzymes in humans, several ofwhich are tumor suppressors. The degradation products of hyaluronicacid, the oligosaccharides and the very-low molecular weight hyaluronicacid, exhibit pro-angiogenic properties. In addition, recent studiesshow that hyaluronan fragments, but not the native high molecular massof hyaluronan, can induce inflammatory responses in macrophages anddendritic cells in tissue injury. Hyaluronic acid is well suited tobiological applications targeting the skin. Due to its highbiocompatibility, it is used to stimulate tissue regeneration. Studieshave shown hyaluronic acid appearing in the early stages of healing tophysically create room for white blood cells that mediate the immuneresponse. It is used in the synthesis of biological scaffolds for woundhealing applications and in wrinkle treatment. A suitable range ofconcentration over which hyaluronic acid can be used in the presentcomposition is from about 0.001% to about 3%.

Glucosamine: is one of the most abundant monosaccharides in humantissues and a precursor in the biological synthesis of glycosilatedproteins and lipids. It is commonly used in the treatment ofosteoarthritis. The common form of glucosamine used is its sulfate salt.Glucosanune shows a number of effects including an anti-inflammatoryactivity, stimulation of the synthesis of proteoglycans and thesynthesis of proteolytic enzymes. A suitable range of concentration overwhich glucosamine can be used in the present composition is from about0.01% to about 3%.

Allantoin: is a diureide of glyosilic acid. It has keratolytic effect,increases the water content of the extracellular matrix, enhances thedesquamation of the upper layers of dead (apoptotic) skin cells, andpromotes skin proliferation and wound healing.

Also, saffron can act as both a chromophore and a healing factor. Otherhealing agents can also be included such as growth factors.

(e) Antimicrobials

Antimicrobials kill microbes or inhibit their growth or accumulation.Exemplary antimicrobials (or antimicrobial agent) are recited in U.S.Patent Application Publications 20040009227 and 20110081530. Suitableantimicrobials for use in the methods of the present disclosure include,but not limited to, phenolic and chlorinated phenolic and chlorinatedphenolic compounds, resorcinol and its derivatives, bisphenoliccompounds, benzoic esters (parabens), halogenated carbonilides,polymeric antimicrobial agents, thazolines, trichloromethylthioimides,natural antimicrobial agents (also referred to as “natural essentialoils”), metal salts, and broad-spectrum antibiotics.

Specific phenolic and chlorinated phenolic antimicrobial agents that canbe used in the disclosure include, but are not limited to phenol,2-methyl phenol, 3-methyl phenol; 4-methylphenol; 4-ethyl phenol;2,4-dimethyl phenol; 2,5-dimethyl phenol; 3,4-dimethyl phenol;2,6-dimethyl phenol; 4-n-propyl phenol; 4-n-butyl phenol; 4-n-amylphenol; 4-tert-amyl phenol; 4-n-hexyl phenol; 4-n-heptyl phenol; mono-and poly-alkyl and aromatic halophenols; p-chlorophenyl; methylp-chlorophenol; ethyl p-chlorophenol; n-propyl p-chlorophenol; n-butylp-chlorophenol; n-amyl p-chlorophenol; sec-amyl p-chlorophenol; n-hexylp-chlorophenol; cyclohexyl p-chlorophenol; n-heptyl p-chlorophenol,n-octyl, p-chlorophenol; o-chlorophenol; methyl o-chlorophenol; ethylo-chlorophenol; n-propyl o-chlorophenol; n-butyl o-chlorophenol; n-amylo-chlorophenol, tert-amyl o-chlorophenol; n-hexyl o-chlorophenol;n-heptyl o-chlorophenol; o-benzyl p-chlorophenol; o-benxyl-m-methylp-chlorophenol; o-benzyl-m,m-dimethyl p-chlorophenol; o-phenylethylp-chlorophenol; o-phenylethyl-m-methyl p-chlorophenol, 3-methylp-chlorophenol 3,5-dimethyl p-chlorophenol, 6-ethyl-3-methylp-chlorophenol, 6-n-propyl-3-methyl p-chlorophenol;6-iso-propyl-3-methyl p-chlorophenol; 2-ethyl-3,5-dimethylp-chlorophenol; 6-sec-butyl-3-methyl p-chlorophenol,2-iso-propyl-3,5-dimethyl p-chlorophenol; 6-diethylmethyl-3-methylp-chlorophenol; 6-iso-propyl-2-ethyl-3-methyl p-chlorophenol;2-sec-amyl-3,5-dimethyl p-chlorophenol; 2-diethylmethyl-3,5-dimethylp-chlorophenol, 6-sec-octyl-3-methyl p-chlorophenol; p-chloro-m-cresolp-bromophenol; methyl p-bromophenol; ethyl p-bromophenol; n-propylp-bromophenol; n-butyl p-bromophenol; n-amyl p-bromophenol; sec-amylp-bromophenol; n-hexyl p-bromophenol; cyclohexyl p-bromophenol;o-bromophenol; tert-amyl o-bromophenol; n-hexyl o-bromophenol;n-propyl-m,m-dimethyl o-bromophenol; 2-phenyl phenol; 4-chloro-2-methylphenol; 4-chloro-3-methyl phenol; 4-chloro-3,5-dimethyl phenol;2,4-dichloro-3,5-dimethylphenol; 3,4,5,6-tetabromo-2-methylphenol-;5-methyl-2-pentylphenol; 4-isopropyl-3-methylphenol;para-chloro-metaxylenol (PCMX); chlorothymol; phenoxyethanol;phenoxyisopropanol; and 5-chloro-2-hydroxydiphenylmethane.

Resorcinol and its derivatives can also be used as antimicrobial agents.Specific resorcinol derivatives include, but are not limited to: methylresorcinol; ethyl resorcinol; n-propyl resorcinol; n-butyl resorcinol;n-amyl resorcinol; n-hexyl resorcinol; n-heptyl resorcinol; n-octylresorcinol; n-nonyl resorcinol; phenyl resorcinol; benzyl resorcinol;phenylethyl resorcinol; phenylpropyl resorcinol; p-chlorobenzylresorcinol; 5-chloro-2,4-dihydroxydiphenyl methane,4′-chloro-2,4-dihydroxydiphenyl methane, 5-bromo-2,4-dihydroxy diphenylmethane, and 4′-bromo-2,4-dihydroxydiphenyl methane.

Specific bisphenolic antimicrobial agents that can be used in thedisclosure include, but are not limited to: 2,2′-methylenebis-(4-chlorophenol); 2,4,4′-trichloro-2′-hydroxy-diphenyl ether, whichis sold by Ciba Geigy, Florham Park, N.J. under the tradenameTriclosan®; 2,2′-methylene bis (3,4,6-trichlorophenol); 2,2′-methylenebis-(4-chloro-6-bromophenol); bis-(2-hydroxy-3,5-dichlorop-henyl)sulphide, and bis-(2-hydroxy-5-chlorobenzyl)sulphide.

Specific benzoic esters (parabens) that can be used in the disclosureinclude, but are not limited to: methylparaben; propylparaben;butylparaben; ethylparaben; isopropylparaben; isobutylparaben;benzylparaben; sodium methylparaben, and sodium propylparaben.

Specific halogenated carbanilides that can be used in the disclosureinclude, but are not limited to: 3,4,4′-trichlorocarbanilides, such as3-(4-chlorophenyl)-1-(3,4-dichlorphenyl)urea sold under the tradenameTriclocarban® by Ciba-Geigy, Florham Park, N.J.;3-trifluoromethyl-4,4′-dichlorocarbanilide; and3,3′,4-trichlorocarbanilide.

Specific polymeric antimicrobial agents that can be used in thedisclosure include, but are not limited to: polyhexamethylene biguanidehydrochloride; and poly(iminoimidocarbonyl iminoimidocarbonyliminohexamethylene hydrochloride), which is sold under the tradenameVantocil® IB.

Specific thazolines that can be used in the disclosure include, bot arenot limited to that sold under the tradename Micro-Check®; and2-n-octyl-4-isothiazolin-3-one, which is sold under the tradenameVinyzene® IT-3000 DIDP.

Specific trichloromethylthioimides that can be used in the disclosureinclude, but are not limited to: N-(trichloromethylthio)phthalimide,which is sold under the tradename Fungitrol®; andN-trichloromethylthio-4-cyclohexene-1,2-dicarboximide, which is soldunder the tradename Vancide®.

Specific natural antimicrobial agents that can be used in the disclosureinclude, but are not limited to, oils of: anise; lemon; orange;rosemary; wintergreen; thyme; lavender; cloves; hops; tea tree;citronella, wheat; barley; lemongrass; cedar leaf; cedarwood; cinnamon;fleagrass; geranium, sandalwood; violet; cranberry; eucalyptus; vervain;peppermint; gum benzoin; basil; fennel; fir; balsam; menthol; ocmeaoriganuin; hydastis; carradensis; Berberidaceac daceae; Ratanhiae longa,and Curcuma longa. Also included in this class of natural antimicrobialagents are the key chemical components of the plant oils which have beenfound to provide antimicrobial benefit. These chemicals include, but arenot limited to: anethol; catechole; camphene; thymol; eugenol;eucalyptol; ferulic acid; famesol; hinokitiol; tropolone; limonene;menthol; methyl salicylate; carvacol; terpineol; verbenone; berberine;ratanhiae extract; caryophellene oxide; citronellic acid; curcumin;nerolidol; and geraniol.

Specific metal salts that can be used in the disclosure include, but arenot limited to, salts of metals in groups 3a-5a, 3b-7b, and 8 of theperodic table. Specific examples of metal salts include, but are notlimited to, salts of aluminum; zirconium; zinc, silver; gold, copper;lanthanunt; tin; mercury; bismuth; selenium; strontium; scandium;yttrium; cerium; praseodymiun; neodymium; promethum; samarium; europium;gadolinium; terbium; dysprosium; holmiom; erbium; thalium; ytterbium;lutetium; and mixtures thereof. An example of the metal-ion basedantimicrobial agent is sold under the tradename HealthShield®, and ismanufactured by HealthShield Technology, Wakefield, Mass. [give otherexamples here e.g. smith and nephew]

Specific broad-spectrum antimicrobial agents that can be used in thedisclosure include, but are not limited to, those that are recited inother categories of antimicrobial agents herein.

Additional antinucrobial agents that can be used in the methods of thedisclosure include, but are not limited to: pyrithiones, and inparticular pyrithione-including zinc complexes such as that sold underthe tradename Octopirox®; dimethyidimethylol hydantoin, which is soldunder the tradename Glydant®;methylchloroisothiazolinone/methylisothiazolinone, which is sold underthe tradename Kathon CG®, sodium sulfite; sodium bisulfite;imidazolidinyl urea, which is sold under the tradename Germall 115®;diazolidinyl urea, which is sold under the tradename Germall 11®; benzylalcohol v2-bromo-2-nitropropane-1,3-diol, which is sold under thetradename Bronopol®; formalin or formaldehyde; iodopropenylbutylcarbamate, which is sold under the tradename Polyphase P100®;chloroacetamide; methanamine; methyldibromonitrile glutaronitrile(1,2-dibromo-2.4-dicyanobutane), which is sold under the tradenameTektamer®; glutaraldehyde; 5-bromo-5-nitro-1,3-dioxane, which is soldunder the tradename Bronidox®; phenethyl alcohol; o-phenylphenol/sodiumo-phenylphenol sodium hydroxymethylglycinate, which is sold under thetradename Suttocide A®; polymethoxy bicyclic oxazolidine; which is soldunder the tradename Nuosept C®: dimethoxane; thimersal; dichlorobenzylalcohol; captan; chlorphenenesin; dichloropbene; chlorbutanol; glyceryllaurate; halogenated diphenyl ethers;2,4,4′-trichloro-2′-hydroxy-diphenyl etber, which is sold under thetradename Triclosan® and is available from Ciba-Geigy, Florham Park,N.J.; and 2,2′-dihydroxy-5,5′-dibromo-diphenyl ether.

Additional antimicrobial agents that can be used in the methods of thedisclosure inclode those disclosed by U.S. Pat. Nos. 3,141,321;4,402,959; 4,430,381; 4,533,435; 4,625,026; 4,736,467; 4,855,139;5,069,907; 5,091,102; 5,639,464; 5,853,883; 5,854,147; 5,894,042; and5,919,554, and U.S. Pat. Appl. Publ. Nos. 20040009227 and 20110081530.

(D) Collagens and Agents that Promote Collagen Synthesis

Collagen is a fibrous protein produced in dermal fibroblast cells andforming 70% of the dermis. Collagen is responsible for the smoothing andfirming of the skin. Therefore. when the synthesis of collagen isreduced, skin aging will occur, and so the firming and smoothing of theskin will be rapidly reduced. As a result, the skin will be flaccid andwrinkled. On the other hand, when metabolism of collagen is activated bythe stimulation of collagen synthesis in the skin, the components ofdermal matrices will be increased, leading to effects, such as wrinkleimprovement, firmness improvement and skin strengthening. Thus,collagens and agents that promote collagen synthesis may also be usefulin the present disclosure. Agents that promote collagen synthesis (i.e.,pro-collagen synthesis agents) include amino acids, peptides, proteins,lipids, small chemical molecules, natural products and extracts fromnatural products.

For instance, it was discovered that intake of vitanun C, iron, andcollagen can effectively increase the amount of collagen in skin orbone. See, e.g., U.S. Patent Application Publication 20090069217.Examples of the vitamin C include an ascorbic acid derivative such asL-ascorbic acid or sodium L-ascorbate, an ascorbic acid preparationobtained by coating ascorbic acid with an emulsifier or the like, and amixture containing two or more of those vitamin Cs at an arbitrary rate.In addition, natural products containing vitamin C such as acerola andlemon may also be used. Examples of the iron preparation include: aninorganic tron such as ferrous sulfate, sodium ferrous citrate, orferric pyrophosphate; an organic tron such as heme iron, ferritin iron,or lactoferrin iron; and a mixture containing two or more of those ironsat an arbitrary rate. In addition, natural products containing iron suchas spinach or liver may also be used. Moreover, examples of the collageninclude: an extract obtained by treating bone, skin, or the like of amammal such as bovine or swine with an acid or alkaline; a peptideobtained by hydrolyzing the extract with a protease such as pepsine,trypsin, or chymotrypsin; and a mixture containing two or more of thosecollagens at an arbitrary rate. Collagens extracted from plant sourcesmay also be used.

Additional pro-collagen synthesis agents are described, for example, inU.S. Pat. Nos. 7,598,291, 7,722,904, 6,203,805 , 5,529,769, etc. andU.S. Patent Application Publications 20060247313, 20080108681,20110130459, 20090325885, 20110086060, etc.

(4) Methods of Use

The biophotonic compositions of the present disclosure have numeroususes. Without being bound by theory, the biophotonic compositions of thepresent disclosure may promote wound healing or tissue repair. Thebiophotonic compositions of the present disclosure may also be used totreat a skin disorder. The biophotonic compositions of the presentdisclosure may also be used to treat acne. The biophotonic compositionsof the present disclosure may also be used for skin rejuvenation. Thebiophotonic compositions of the present disclosure may also be used fortreating acute inflammation. Therefore, it is an objective of thepresent disclosure to provide a method for providing biophotonic therapyto a wound, where the method promotes wound healing. It is also anobjective of the present disclosure to provide a method for providingbiophotonic therapy to a skin tissue afflicted with acne, wherein themethod is used to treat acne. It is also an objective of the presentdisclosure to provide a method for providing biophotonic therapy to askin tissue afflicted with a skin disorder, wherein the method is usedto treat the skin disorder. It is also an objective of the presentdisclosure to provide a method for providing biophotonic therapy to skintissue, wherein the method is used for promoting skin rejuvenation.

In certain embodiments, the present disclosure provides a method forproviding a biophotonic therapy to a wound, the method comprising:applying (e.g., by topical application) a biophotonic composition of thepresent disclosure to a site of a wound, and illuminating thebiophotonic composition with light having a wavelength that overlapswith an absorption spectrum of the chromophore(s) of the biophotoniccomposition.

In one aspect, the present disclosure provides a method for providingbiophotonic therapy to a wound, comprising: topically applying abiophotonic composition comprising a first chromophore, and illuminatingsaid biophotonic composition with light having a wavelength thatoverlaps with an absorption spectrum of the first chromophore, whereinthe biophotonic composition is substantially resistant to leaching suchthat it limits leaching of the chromophore into the tissue duringtreatment. In some embodiments, less than 30%, 25%, 20%, 15%, 10%, 5%,1%, 0.8%, 0.5% or 0 1% or essentially 0% of the total chromophore amountleaches out of the biophotonic composition into the wound or tissueduring treatment.

In another aspect, the present disclosure provides a method for treatinga wound or providing biophotonic therapy to a wound, comprising:topically applying a biophotonic composition comprising a firstchromophore and a gelling agent to a site of a wound; and illuminatingsaid biophotonic composition with light having a wavelength thatoverlaps with an absorption spectrum of the first chromophore, whereinthe gelling agent blocks substantial leaching of the chromophores intothe site of a wound during treatment. In some embodiments, less than30%, 25%, 20%, 15%, 10%, 5%, 1%, 0.8%, 05% or 01% or essentially 0% ofthe total chromophore amount leaches out of the biophotonic compositioninto the wound or tissue during treatment.

In yet another aspect, the present disclosure provides a method forpromoting skin rejuvenation. In certain embodiments, the presentdisclosure provides a method for providing skin rejuvenation, the methodcomprising: applying (e.g., by topical application) a biophotomiccomposition of the present disclosure to the skin, and illummating thebiophotonic composition with light having a wavelength that overlapswith an absorption spectrum of the chromophore(s) of the biophotoniccomposition.

In other embodiments, the present disclosure provides a method forpromoting skin rejuvenation comprising: topically applying a biophotoniccomposition comprising a first chromophore to skin; and illummating saidbiophotonic composition with light having a wavelength that overlapswith an absorption spectrum of the first chromophore; wherem thebiophotonic composition as substantially resistant to leaching such thatit limits leaching of the chromophore into the skin during treatment. Insome embodiments, less than 30%, 25%, 20%, 15%, 10%, 5%, 1%, 0.8%, 0.5%or 0.1% or essentially 0% of the total chromophore amount leaches out ofthe biophotonic composition into the wound or tissue during treatment.

In another aspect, the present disclosure provides a method forpromoting skin rejuvenation, comprising: topically applying abiophotonic composition comprising a first chromophore and a gellingagent to skin, and illuminating said biophotonic composition with lighthaving a wavelength that overlaps with an absorption spectrum of thefirst chromophore; wherein the biophotonic composition is substantiallyresistant to leaching such that it blocks substantial leaching of thechromophores into the skin during treatment. In some embodiments, lessthan 30%, 25%, 20%, 15%, 10%, 5%, 1%, 0 8%, 0.5% of 0.1% of essentially0% of the total chromophore amount leaches out of the biophotoniccomposition into the skin during treatment.

In yet another aspect, the present disclosure to provide a method forproviding biophotonic therapy to a target skin tissue afflicted with askin disorder. In certain embodiments, the present disclosure provides amethod for providing a biophotonic therapy to a target skin tissue, themethod comprising: applying (e.g., by topical application) a biophotomiccomposition of the present disclosure to a target skin tissue, andilluminating the biophotonic composition with light having a wavelengththat overlaps with an absorption spectrum of the chromophore(s) of thebiophotonic composition.

In other embodiments, the present disclosure provides a method fortreating a skin disorder, comprising: topically applying a biophotoniccomposition to a target skin tissue afflicted with the skin disorder,wherein the biophotonic composition comprises a first chromophore, andilluminating said biophotonic composition with light having a wavelengththat overlaps with an absorption spectrum of the first chromophore;wherein the biophotonic composition as substantially resistant toleaching such that it limits leaching of the chromophore into the skinduring treatment. In some embodiments, less than 30%, 25%, 20%, 15%,10%, 5%, 1%, 0.8%, 0.5% or 0 1% or essentially 0% of the totalchromophore amount leaches out of the biophotonic composition into theskin during treatment.

In another aspect, the present disclosure provides a method for treatinga skin disorder, comprising: topically applying a biophotoniccomposition comprising a first chromophore and a gelling agent to skinafflicted with the skin disorder; and illuminating said biophotoniccomposition with light having a wavelength that overlaps with anabsorption spectrum of the first chromophore; wherein the biophotoniccomposition is substantially resistant to leaching such that it blockssubstantial leaching of the chromophores into the skin during treatment.In some embodiments, less than 30%, 25%, 20%, 15%, 10%, 5%, 1%, 0.8%,0.5% or 0.1% or essentially 0% of the total chromophore amount leachesout of the biophotonic composition into the skin during treatment.

In yet another aspect, the present disclosure to provide a method forproviding biophotonic therapy to a target skin tissue afflicted withacne.In certain embodiments, the present disclosure provides a methodfor providing a biophotonic therapy to a target skin tissue afflictedwith acne, the method comprising: applying (e.g., by topicalapplication) a biophotonic composition of the present disclosure to atarget skin tissue, and illuminating the biophotonic composition withlight having a wavelength that overlaps with an absorption spectrum ofthe chromophore(s) of the biophotonic composition.

In other embodiments, the present disclosure provides a method fortreating acne, comprising: topically applying a biophotonic compositionto a target skin tissue afflicted with acne, wherein the biophotoniccomposition comprises a first chromophore; illuminating said biophotoniccomposition with light having a wavelength that overlaps with anabsorption spectrum of the first chromophore; wherein the biophotoniccomposition is substantially resistant to leaching such that it limitsleaching of the chromophore into tissue during treatment. In someembodiments, less than 30%, 25%, 20%, 15%, 10%, 5%, 1%, 0.8%, 0.5% or0.1% or essentially 0% of the total chromophore amount leaches out ofthe biophotonic composition into the tissue during treatment.

In another aspect, the present disclosure provides a method for treatingacne, comprising: topically applying a biophotonic compositioncomprising a first chromophore to skm afflicted with acne; andilluminating said biophotonic composition with light having a wavelengththat overlaps with an absorption spectrum of the first chromophore;wherein the biophotonic composition is substantially resistant toleaching such that it blocks substantial leaching of the chromophoresinto the skin during treatment. In some embodiments, less than 30%, 25%,20%, 15%, 10%, 5%, 1%, 0.8%, 0.5% or 0.1% or essentially 0% of the totalchromophore amount leaches out of the biophotonic composition into thewound or tissue during treatment.

In other embodiments, the present disclosure provides a method fortreating acute inflammation, comprising: topically applying abiophotonic composition to a target skin tissue with acute inflammation,wherein the biophotonic composition comprises a first chromophore,illuminating said biophotonic composition with light having a wavelengththat overlaps with an absorption spectrum of the first chromophore;wherein the biophotonic composition is substantially resistant toleaching such that it limits leaching of the chromophore into tissueduring treatment. In some embodiments, less than 30%, 25%, 20%, 15%,10%, 5%, 1%, 0 8%, 0.5% or 0 1% or essentially 0% of the totalchromophore amount leaches out of the biophotonic composition into thetissue during treatment.

In another aspect, the present disclosure provides a method for treatingacute inflammation, comprising: topically applying a biophotoniccomposition comprising a first chromophore to skin afflicted with acuteinflammation; and illuminating said biophotonic composition with lighthaving a wavelength that overlaps with an absorption spectrom of thefirst chromophore; wherein the biophotonic composition is substantiallyresistant to leaching such that it blocks substantial leaching of thechromophores into the skin during treatment. In some embodiments, lessthan 30%, 25%, 20%, 15%, 10%, 5%, 1%, 0.8%, 0.5% or 0 1% or essentially0% of the total chromophore amount leaches out of the biophotoniccomposition into the wound or tissue during treatment.

The biophotonic compositions suitable for use in the methods of thepresent disclosure may be selected from any of the embodiments of thebiophotonic compositions described above. For instance, the biophotoniccompositions useful in the method of the present disclosure may comprisea first chromophore that undergoes at least partial photobleaching uponapplication of light. The first chromophore may absorb at a wavelengthof about 200-800 nm, 200-700 nm, 200-600 nm or 200-500 nm. In oneembodiment, the first chromophore absorbs at a wavelength of about200-600 nm. In some embodiments, the first chromophore absorbs light ata wavelength of about 200-300 nm, 250-350 nm, 300-400 nm. 350-450 nm,400-500 nm, 450-650 nm, 600-700 nm, 650-750 nm or 700-800 nm. In otherexamples, suitable biophotonic compositions for the methods of thepresent disclosure may further comprise at least one additionalchromophore (e.g., a second chromophore). The absorption spectrum of thesecond chromophore overlaps at least about 80%, 50%, 40%, 30%, or 20%with the emission spectrum of the first chromophore. In someembodiments, the first chromophore bas an emission spectrum thatoverlaps at least 1-10%, 5-15%, 10-20%, 15-25%, 20-30%, 25-35%, 30-40%,35-45%, 50-60%, 55-65% or 60-70% with an absorption spectrum of thesecond chromophore.

Illumination of the biophotonic composition with light may cause atransfer of energy from the first chromophore to the second chromophore.Subsequently, the second chromophore may emit energy as fluorescenceand/or generate reactive oxygen species. In certain embodiments of themethods the present disclosure, energy transfer caused by theapplication of light is not accompanied by concomitant generation ofheat, or does not result in tissue damage.

The biophotonic compositions useful for the present methods comprise agelling agent. The gelling agent may include, but is not limited to,lipids such as glycerin, glycols such as propylene glycol, hyaluronicacid, glucosamine sulfate, cellulose derivatives (hydroxypropylmethylcellulose, hydroxyethyl cellulose, hydroxypropyl cellulose,methylcellulose and the like), noncellulose polysaccharides(galactomannans, guar gum, carob gum, gum arabic, sterculia gum, agar,alginates and the like) and acrylic acid polymers.

When the method involves a biophotonic composition having at least twochromophores, the first chromophore is present in an amount of about 001-40% per weight of the composition, and the second chromophore ispresent in an amount of about 0.001-40% per weight of the composition.In certam embodiments, the first chromophore is present in an amount ofabout 0.01-1%, 0.5-2%, 1-5%, 2.5-7.5%, 5-10%, 7.5-12.5%, 10-15%,12.5-17.5%, 15-20%, 17.5-22.5%, 20-25%, 22.5-27.5%, 25-30%, 27.5-32.5%,30-35%, 32.5-37.5%, or 35-40% per weight of the composition. In certainembodiments, the second chromophore is present in an amount of about0.001-1%, 0.5-2%, 1-5%, 2.5-7.5%, 5-10%, 7.5-12.5%, 10-15%, 12.5-17.5%,15-20%, 17.5-22.5%, 20-25%, 22.5-27 5%, 25-30%, 27.5-32.5%, 30-35%,32.5-37.5%, of 35-40% per weight of the composition. In certainembodiments, the total weight per weight of chromophore or combinationof chromophores may be in the amount of about 0.01-1%, 0.5-2%, 1-5%,2.5-7 5%, 5-10%, 7.5-12.5%, 10-15%, 12.5-17.5%, 15-20%, 17.5-22.5%,20-25%, 22.5-27.5%, 25-30%, 27.5-32 5%, 30-35%, 32.5-37.5%, or 35-40.05%per weight of the composition.

In the methods of the present disclosure, any source of actinic lightcan be used. Any type of halogen, LED or plasma arc lamp or laser may besuitable. The primary characteristic of suitable sources of actiniclight will be that they emit light in a wavelength for wavelengths)appropriate for activating the one or more photoactivators present inthe composition. In one embodiment, an argon laser is used. In anotherembodiment, a potassium-titanyl phosphate (KTP) laser (e.g. aGreenLight™ laser) is used. In another embodiment, sunlight may be used.In yet another embodiment, a LED photocuring device is the source of theactinic light. In yet another embodiment, the source of the actiniclight is a source of light having a wavelength between about 200 to 800nm. In another embodiment, the source of the actinic light is a sourceof visible light having a wavelength between about 400 and 600 nm.Furthermore, the source of actinic light should have a suitable powerdensity. Suitable power density for non-collimated light sources (LED,halogen or plasma lamps) are in the range from about 1 mW/cm² to about200 mW/cm². Suitable power density for laser light sources are in therange from about 0.5 mW/cm² to about 0.8 mW/cm².

In some embodiments of the methods of the present disclosure, the lighthas an energy at the subject's skin, wound or mucosa surface of betweenabout 1 mW/cm² and about 500 mW/cm², 1-300 mW/cm², or 1-200 mW/cm²,wherein the energy applied depends at least on the condition beingtreated, the wavelength of the light, the distance of the subject's skinfrom the light source, and the thickness of the biophotonic compsoition.In certain embodiments, the light at the subject's skin is between about1-40 mW/cm², or 20-60 mW/cm², or 40-80 mW/cm², or 60-100 mW/cm², or80-120 mW/cm², or 100-140 mW/cm², or 120-160 mW/cm², or 140-180 mW/cm²,or 160-200 mW/cm², or 110-240 mW/cm², or 110-150 mW/cm², or 190-240mW/cm².

In some embodiments, a mobile device can be used to activate embodimentsof the biophotonic composition of the present disclosure, wherem themobile device can emit light having an emission spectra which overlapsan absorption spectra of the chromophore in the biophotonic composition.The mobile device can have a display screen through which the light isemitted and/or the mobile device can emit light from a flashlight whichcan photoactivate the biophotonic composition.

In some embodiments, a display screen on a television or a computermonitor can be used to activate the biophotonic composition, wherein thedisplay screen can emnt light having an emission spectra which overlapsan absorption spectra of a photoactive agent in the photoactivatablecomposition.

In certain embodiments, the first and/or the second chromophore (whenpresent) can be photoactivated by ambient light which may originate fromthe sun or other light sources. Ambient light can be considered to be ageneral illumination that comes from all directions in a room that hasno visible source. In certain embodiments, the first and/or the secondchromophore (when present) can be photoactivated by light in the visiblerange of the electromagnetic spectrum. Exposure times to ambient lightmay be longer than that to direct light.

In certain embodiments, different sources of light can be used toactivate the biophotonic compositions, such as a combination of ambientlight and direct LED light.

The duration of the exposure to actinic light required will be dependenton the surface of the treated area, the type of lesion, trauma or injurythat is being treated, the power density, wavelength and bandwidth ofthe light source, the thickness of the biophotonic composition, and thetreatment distance from the light source. The illumination of thetreated area by fluorescence may take place within seconds or evenfragment of seconds, but a prolonged exposure period is beneficial toexploit the synergistic effects of the absorbed, reflected and reemittedlight on the composition of the present disclosure and its interactionwith the fissoe being treated. In one embodiment, the time of exposureto actinic light of the tissue, skin of wound on which the biophotoniccomposition has been applied is a period between 1 minute and 5 minutes.In another embodiment, the time of exposure to actinic light of thetissue, skin or wound on which the biophotonic composition has beenapplied is a period between 1 minute and 5 minutes. In some otherembodiments, the biophotonic composition is illuminated for a periodbetween 1 minute and 3 minutes. In certain embodiments, light is appliedfor a period of 1-30 seconds, 15-45 seconds, 30-60 seconds, 0.75-1.5minutes, 1-2 minutes, 1.5-2.5 minutes, 2-3 minutes, 2.5-3.5 minutes, 3-4minutes, 3.5-4.5 minutes, 4-5 minutes, 5-10 minutes, 10-15 minutes,15-20 minutes, 20-25 minutes, or 20-30 minutes. In yet anotherembodiment, the source of actinic light is in continuous motion over thetreated area for the appropriate time of exposure. In yet anotherembodiment, multiple applications of the biophotonic composition andactinic light are performed. In some embodiments, the tissue, skin orwound is exposed to actinic light at least two, three, four, five or sixtimes. In some embodiments, a fresh application of the biophotoniccomposition is applied before exposure to actinic light.

In the methods of the present disclosure, the biophotonic compositionmay be optionally removed from the site of treatment followingapplication of light. In certain embodiments, the biophotoniccomposition is left on the treatment site for more than 30 minutes, morethan one hour, more than 2 hours, more than 3 hours. It can beilluminated with ambient light. To prevent drying, the composition canbe covered with a transparent of translucent cover such as a polymerfilm, or an opaque cover which can be removed before illumination.

(5) Wounds and Wound Healing

The biophotonic compositions and methods of the present disclosure maybe used to treat wounds and promote wound healing. Wounds that may betreated by the biophotonic compositions and methods of the presentdisclosure include, for example, injuries to the skin and subcutaneoustissue initiated in different ways (e.g., pressure ulcers from extendedbed rest, wounds induced by trauma, wounds induced by conditions such asperiodontitis) and with varying characteristics. In certain embodiments,the present disclosure provides biophotonic compositions and methods fortreating and/or promoting the healing of, for example, burns, incisions,excisions, lacerations, abrasions, puncture or penetrating wounds,surgical wounds, contusions, hematomas, crushing injuries, sores andulcers.

Biophotonic compositions and methods of the present disclosure may beused to treat and/or promote the healing of chronic cutaneous ulcers orwounds, which are wounds that have failed to proceed through an orderlyand timely series of events to produce a durable structural, functional,and cosmetic closure. The vast majority of chronic wounds can beclassified into three categories based on their etiology: pressureulcers, neuropathic (diabetic foot) ulcers and vascular (venous orarterial) ulcers.

In certain other embodiments, the present disclosure providesbiophotonic compositions and methods for treating and/or promotinghealing, Grade I-IV ulcers. In certain embodiments, the applicationprovides compositions suitable for use with Grade II ulcers inparticular. Ulcers may be classified into one of four grades dependingon the depth of the wound: i) Grade I: wounds limited to the epithelium;ii) Grade II: wounds extending into the dermis: iii) Grade III: woundsextending into the subcutaneous tissue: and iv) Grade IV (orfull-thickness wounds): wounds wherein bones are exposed (e.g., a bonypressure point such as the greater trochanter or the sacrum).

For example, the present disclosure provides biophotonic compositionsand methods for treating and/or promoting healing of a diabetic ulcer.Diabetic patients are prone to foot and other ulcerations due to bothneurologic and vascular complications. Peripheral neuropathy can causealtered or complete loss of sensation in the foot and/or leg. Diabeticpatients with advanced neuropathy lose all ability for sharp-dulldiscrimination. Any cuts or trauma to the foot may go completelyunnoticed for days or weeks in a patient with neuropathy. A patient withadvanced neuropathy loses the ability to sense a sustained pressureinsult, as a result, tissue ischemia and necrosis may occur leading tofor example, plantar ulcerations. Microvascular disease is one of thesignificant complications for diabetics which may also lead toulcerations. In certain embodiments, compositions and methods oftreating a chronic wound are provided here in, where the chronic woundis characterized by diabetic foot ulcers and/or ulcerations due toneurologic and/or vascular complications of diabetes.

In other examples, the present disclosure provides biophotoniccompositions and methods for treating and/or promoting healing of apressure ulcer. Pressure ulcer includes bed sores, decubitus ulcers andischial tuberosity ulcers and can cause considerable pain and discomfortto a patient. A pressure ulcer can occur as a result of a prolongedpressure applied to the skin. Thus, pressure can be exerted on the skinof a patient due to the weight or mass of an individual. A pressureulcer can develop when blood supply to an area of the skin is obstructedor cut off for more than two or three hours. The affected skin area canturns red, becomes painful and can become necrotic. If untreated, theskin breaks open and can become infected. An ulcer sore is therefore askin ulcer that occurs m an area of the skin that is under pressure frome.g. lying in bed, sitting in a wheelchair, and/or wearing a cast for aprolonged period of time. Pressure ulcer can occur when a person isbedridden, unconscious, unable to sense pain, or immobile. Pressureulcer often occur in boney prominences of the body such as the buttocksarea (on the sacrum or iliac crest), or on the heels of foot.

In other examples, the present disclosure provides biophotoniccompositions and methods for treating and/or promoting healing of acutewounds.

Additional types of wound that can be treated by the biophotomecompositions and methods of the present disclosure include thosedisclosed by U.S. Pat. Appl. Publ. No. 20090220450, which isincorporated herein by reference.

Wound healing in adult tissues is a complicated reparative process. Forexample, the healing process for skin involves the recruitment of avariety of specialized cells to the site of the wound, extracellularmatrix and basement membrane deposition, angiogenesis, selectiveprotease activity and re-epithelialization.

There are three distinct phases in the wound healing process. First, inthe inflammatory phase, which typically occurs from the moment a woundoccurs until the first two to five days, platelets aggregate to depositgranules, promoting the deposit of fibrin and stimulating the release ofgrowth factors. Leukocytes migrate to the wound site and begin to digestand transport debris away from the wound. During this inflammatoryphase, monocytes are also converted to macrophages, which release growthfactors for stimulating angiogenesis and the production of fibroblasts.

Second, in the proliferative phase, which typically occurs from two daysto three weeks, granulation tissue forms, and epithelialization andcontraction begin. Fibroblasts, which are key cell types in this phase,proliferate and synthesize collagen to fill the wound and provide astrong matrix on which epithelial cells grow. As fibroblasts producecollagen, vascularization extends from nearby vessels, resulting ingranulation tissue. Granulation tissue typically grows from the base ofthe wound. Epithelialization involves the migration of epithelial cellsfrom the wound surfaces to seal the wound. Epithelial cells are drivenby the need to contact cells of like type and are guided by a network offibrin strands that function as a grid over which these cells migrate.Contractile cells called myofibroblasts appear in wounds, and aid inwound closore. These cells exhibit collagen synthesis and contractility,and are common in granulating wounds.

Third, in the remodeling phase, the final phase of wound healing whichcan take place from three weeks up to several years, collagen in thescar undergoes repeated degradation and re-synthesis. During this phase,the tensile strength of the newly formed skin increases.

However, as the rate of wound healing increases, there is often anassociated increase in scar formation. Scarring is a consequence of thehealing process in most adult animal and human tissues. Scar tissue isnot identical to the tissue which it replaces, as it is usually ofinferior functional quality. The types of scars include, but are notlimited to, atrophic, hypertrophic and keloidal scars, as well as scarcontractures. Atrophic scars are flat and depressed below thesurrounding skin as a valley or hole. Hypertrophic scars are elevatedscars that remain within the boundaries of the original lesion, andoften contain excessive collagen arranged in an abnormal pattern.Keloidal scars are elevated scars that spread beyond the margins of theoriginal wound and invade the surrounding normal skin in a way that issite specific, and often contain whorls of collagen arranged in anabnormal fashion.

In contrast, normal skin consists of collagen fibers arranged in abasket-weave pattern, which contributes to both the strength andelasticity of the dermis. Thus, to achieve a smoother wound healingprocess, an approach is needed that not only stimulates collagenproduction, but also does so in a way that reduces scar formation.

The biophotonic compositions and methods of the present disclosurepromote the wound healing by promoting the formation of substantiallyuniform epithelialization; promoting collagen synthesis, promotingcontrolled contraction, and/or by reducing the formation of scar tissue.In certain embodiments, the biophotonic compositions and methods of thepresent disclosure may promote wound healing by promoting the formationof substantially uniform epithelialization. In some embodiments, thebiophotonic compositions and methods of the present disclosure promotecollagen synthesis. In some other embodiments, the biophotoniccompositions and methods of the present disclosure promote controlledcontraction. In certain embodiments, the biophotonic compositions andmethods of the present disclosure promote wound healing, for example, byreducing the formation of scar tissue or by speeding up the woundclosure process. In certam embodiments, the biophotonic compositions andmethods of the present disclosure promote wound healing, for example, byreducing inflammation. In certain embodiments, the biophotoniccomposition can be used following wound closure to optimize scarrevision. In this case, the biophotonic composition may be apphed atregular intervals such as once a week, or at an interval deemedappropriate by the physician.

The biophotonic composition may be soaked into a woven or non-wovenmaterial or a sponge and applied as a wound dressing. A light source,such as LEDs or waveguides, may be provided within or adjacent the wounddressing or the composition to illuminate the composition. Thewaveguides can be optical fibres which can transmit light, not only fromtheir ends, but also from their body. For example, made of polycarbonateor polymethylmethacrylate.

Adjunct therapies which may be topical or systemie such as antibiotictreatment may also be used. Negative pressure assisted wound closure canalso be used to assist wound closure and/or to remove the composition.

(6) Acne and Acne Scars

The biophotonic compositions and methods of the present disclosure maybe used to treat acne. As used herein, “acne” means a disorder of theskin caused by inflammation of skin glands or hair follicles. Thebiophotonic compositions and methods of the disclosure can be used totreat acne at early pre-emergent stages or later stages where lesionsfrom acne are visible. Mild, moderate and severe acne can be treatedwith embodiments of the biophotonic compositions and methods. Earlypre-emergent stages of acne usually begin with an excessive secretion ofsebum or dermal oil from the sebaceous glands located in thepilosebaceous apparatus. Sebum reaches the skin surface through the ductof the hair follicle. The presence of excessive amounts of sebum in theduct and on the skin tends to obstruct or stagnate the normal flow ofsebum from the follicular duct, thus producing a thickening andsolidification of the sebum to create a solid plug known as a comedone.In the normal sequence of developing acne. hyperkeratinazation of thefolhcular opening is stimulated, thus completing blocking of the duct.The usual results are papules, pustules, or cysta, often contaminatedwith bacteria, which cause secondary infections. Acne is characterizedparticularly by the presence of comedones, inflammatory papules, orcysts. The appearance of acne may range from slight skin irritation topitting and even the development of disfiguring scars. Accordingly, thebiophotonic compositions and methods of the present disclosure can beused to treat one or more of skin irritation, pitting, development ofscars, comedones, inflammatory papules, cysts, hyperkeratinazation, andthickening and hardening of sebum associated with acne.

The composition may be soaked into or applied to a woven or non-wovenmaterial or a sponge and applied as a mask to body parts such as theface, body, arms, legs etc. A light source, such as LEDs or waveguides,may be provided within or adjacent the mask or the composition toilluminate the composition. The waveguides can be optical fibres whichcan transmit light, not only from their ends, but also from their body.For example, made of polycarbonate or polymethylmethacrylate.

The biophotonic compositions and methods of the present disclosure maybe used to treat various types of acne. Some types of acne include, forexample, acne vulgaris, cystic acne, acne atrophica, bromide acne,chlorine acne, acne conglobata, acne cosmetica, acne detergicans,epidemic acne, acne estivalis, acne fulminans, halogen acne, acneindurata, iodide acne, acne keloid, acne mechanica, acne papulosa,pomade acne, premenstral acne, acne postulosa, acne scorbutica, acnescrofalosorum, acne urticata, acne varioliformis, acne venenata,propionic acne, acne excoriee, gram negative acne, steroid acne, andnodulocystic acne.

(7) Skin Aging and Rejuvenation

The dermis is the second layer of skin, containing the structuralelements of the skin, the connective tissue. There are various types ofconnective tissue with different functions. Elastin fibers give the skinits elasticity, and collagen gives the skin its strength.

The junction between the dermis and the epidermis is an importantstructure. The dermal-epidermal junction interlocks forming finger-likeepidermal ridges. The cells of the epidermis receive their nutrientsfrom the blood vessels in the dermis. The epidermal ridges increase thesurface area of the epidermis that is exposed to these blood vessels andthe needed nutrients.

The aging of skin comes with significant physiological changes to theskin. The generation of new skin cells slows down, and the epidermalridges of the dermal-epidermal junction flatten out. While the number ofelastin fibers increases, their structure and coherence decrease. Alsothe amount of collagen and the thickness of the dermis decrease with theageing of the skin.

Collagen is a major component of the skin's extracellular matrix,providing a structural framework. During the aging process, the decreaseof collagen synthesis and insolubilization of collagen fibers contributeto a thinning of the dermis and loss of the skin's biomechanicalproperties.

The physiological changes to the skin result in noticeable agingsymptoms often referred to as chronological-, intrinsic- andphoto-ageing. The skin becomes drier, roughness and scaling increase,the appearance becomes duller, and most obviously fine lines andwrinkles appear. Other symptoms or signs of skin aging include, but arenot limited to, thinning and transparent skin, loss of underlying fat(leading to hollowed cheeks and eve sockets as well as noticeable lossof firmness on the hands and neck), bone loss (such that bones shrinkaway from the skin due to bone loss, which causes sagging skin), dryskin (which might itch), inability to sweat sufficiently to cool theskin, unwanted facial hair, freckles, age spots, spider veins, rough andleathery skin, fine wrinkles that disappear when stretched, loose skin,a blotchy complexion.

The dermal-epidermal junction is a basement membrane that separates thekeratmocytes in the epidermis from the extracellular matrix, which lesbelow in the dermis. This membrane consists of two layers: the basallamina in contact with the keratinocytes, and the underlying reticularlamina in contact with the extracellular matrix. The basal lamina isrich in collagen type IV and lananin, molecules that play a role inproviding a structural network and bioadhesive properties for cellattachment.

Laminin is a glycoprotein that only exists in basement membranes. It iscomposed of three polypeptide chains (alpha, beta and gamma) arranged inthe shape of an asymmetric cross and held together by disulfide bonds.The three chains exist as different subtypes which result in twelvedifferent isoforms for laminin, including Laminin-1 and Laminin-5.

The dermis is anchored to hemidesmosomes, specific junction pointslocated on the keratmocytes, which consist of α-integrins and otherproteins, at the basal membrane keratinocytes by type VII collagenfibrils. Laminins, and particularly Laminin-5, constitute the realanchor point between hemidesmosomal transmembrane proteins in basalkeratinocytes and type VII collagen.

Lammin-5 synthesis and type VII collagen expression have been proven todecrease in aged skin. This causes a loss of contact between dermis andepidermis, and results in the skin losing elasticity and becoming saggy.

Recently another type of wrinkles generally referred to as expressionwrinkles, got general recognition. These wrinkles require loss ofresilience, particularly in the dermis, because of which the skin is nolonger able to resume its original state when facial muscles whichproduce facial expressions exert stress on the skin, resulting inexpression wrinkles.

The compositions and methods of the present disclosure promote skinrejuvenation. In certain embodiments, the compositions and methods ofthe present disclosure promote collagen synthesis. In certain otherembodiments, the compositions and methods of the present disclosure mayreduce, dinunish, retard or even reverse one or more signs of skin agingincluding, but not limited to, appearance of fine lines or wrinkles,thin and transparent skin, loss of underlying fat (leading to hollowedcheeks and eye sockets as well as noticeable loss of firmness on thehands and neck), bone loss (such that bones shrink away from the skindue to bone loss, which causes sagging skin), dry skin (which mightitch), inability to sweat sufficiently to cool the skin, unwanted facialhair, freckles, age spots, spider veins, rough and leathery skin, finewrinkles that disappear when stretched, loose skin, or a blotchycomplexion. In certain embodiments, the compositions and methods of thepresent disclosure may indoce a reduction in pore size, enhancesculpturing of skin subsections, and/or enhance skin translucence.

(8) Skin Disorders

The biophotonic compositions and methods of the present disclosure maybe used to treat skin disorders that include, but are not limited to,erythema, telangiectasia, actinic telangiectasia, psoriasis, skincancer, pemphigus, sunburn, dermatitis, eczema, rashes, impetigo, lichensimplex chronicus, rhinophyma, perioral dermatitis, pseudofolliculitisbarbae, drug eruptions, erythema multiforme, erythema nodosum, granulomaannulare, actinic keratosis, purpura, alopecia areata aphthousstomatitis, drug eruptions, dry skin, chapping, xerosis, ichthyosisvulgaris, fungal infections, parasitic infection, herpes simplex,intertrigo, keloids, keratoses, milia, moluscum contagiosum, pityriasisrosea, pruritus, urticaria, and vascular tumors and malformations.Dermatitis includes contact dermatitis, atopic dermatitis, seborrheicdermatitis, nummular dermatitis, generalized exfoliative dermatitis, andstatis dermatitis. Skin cancers include melanoma, basal cell carcinoma,and squamous cell carcinoma.

Some skin disorders present various symptoms including redness,flushing, burning, scaling, pimples, papules, pustules, comedones,macules, nodules, vesicles, blisters, telangiectasia, spider veins,sores, surface irritations or pain, itching, inflammation, red, purple.or blue patches or discolorations, moles, and/or tumors. Accordingly,the biophotonic compositions and methods of the present disclosure canbe used to treat redness, flushing, burning, scaling, pimples, papules,pustules, comedones, macales, nodules, vesicles, blisters,telangiectasia, spider veins, sores, surface irritations or pain,itching, acute inflanumnation, red, purple, or blue patches ordiscolorations, moles, and/or tumors. Acute inflammation can presentitself as pain, heat, redness, swelling and loss of function. Itincludes those seen in allergic reactions such as insect bites e.g.;mosquito, bees, wasps, poison ivy, post-ablative treatment.

The composition may be soaked into or applied to a woven or non-wovenmaterial or a sponge and applied as a mask to body parts to treat skindisorders. A light source, such as LEDs or waveguides, may be providedwithin or adjacent the mask or the composition to illuminate thecomposition. The waveguides can be optical fibres which can transmitlight, not only from their ends, but also from their body. For example,made of polycarbonate or polymethylmethacry late.

(9) Kits

The present disclosure also provides kits for preparing and/or applyingany of the compositions of the present disclosure. The kit may include abiophotonic topical composition comprising at least a first chromophorein a gelling agent. The composition may include an oxygen-releasingagent present in amount about 0.01%-40%, 0.01%-1.0%, 0.5%-100%, 5%-15%,10%-20%, 15%-25%, 20%-30%, 15.0%-25%, 20% -30%, 25%-35%, or 30%-40% byweight to weight of the composition. The chromophore may be present inan amount of about 0.001-0.1%, 0.05-1%, 0.5-2%, 1-5%, 2.5-7.5%, 5-10%,7.5-12.5%, 10-15%, 12.5-17.5%, 15-20%, 17.5-22.5%, 20-25%, 22.5-27.5%,25-30%. 27.5-32.5%, 30-35%, 32.5-37.5%, or 35-40% per weight of thecomposition. In embodiments where the composition comprises more thanone chromophore, the first chromophore may be present in an amount ofabout 0.01-40% per weight of the composition, and a second chromophoremay be present in an amount of about 0.01-40% per weight of thecomposition. In certain embodiments, the first chromophore is present inan amount of about 0.001-0.1%, 0.05-1%, 0.5-2%, 1-5%, 2.5-7.5%, 5-10%,7.5-12.5%. 10-15%. 12.5-17.5%, 15-20%, 17.5-22.5%, 20-25%, 22.5-27.5%,25-30%, 27.5-32.5%, 30-35%, 32.5-37.5%, or 35-40% per weight of thecomposition. In certain embodiments, the second chromophore is presentin an amount of about 0 001-0.1%, 0.05-1%, 0.5-2%, 1-5%, 2.5-7.5%,5-10%, 7.5S-12.5%, 10-15%, 12.5-17.5%, 15-20%, 17.5-22.5%, 20-25%,22.5-27.5%. 25-30%, 27.5-32.5%, 30-35%, 32.5-37.5%, or 35-40% per weightof the composition. In certain embodiments, the amount of chromophore orcombination of chromophores may be in the amount of about 0.05-40.05%per weight of the composition. In certain embodiments, the amount ofchromophore of combination of chromophores may be in the amount of about0.001-0.1%, 0.05.1%, 0.5-2%, 1-5%, 2.5-7.5%, 5-10%, 7.5-12.5%, 10-15%,12.5-17.5%, 15-20%, 17.5-22.5%, 20-25%, 22.5-27.5%, 25-30%, 27.5-32.5%,30-35%, 32.5-37.5%, or 35-40.05% per weight of the composition.

In some embodiments, the kit includes more than one composition, forexample, a first and a second composition. The first composition mayinclude the oxygen-releasing agent and the second composition mayinclude the first chromophore in the gelling agent. The firstchromophore may have an emission wavelength between about 400 nm andabout 570 nm. The oxygen releasing agent may be present in the firstcomposition in an amount of about 0.01%-1.0%, 0.5%-10.0%, 5%-15%,10%-20%, 15%-25%, 20%-30%, 15.0%-25%, 20%-30%, 25%-35%, 30%-40% or35%-45% by weight to weight of the first composition. The chromophoremay be present in the second composition in an amount of about0.001-0.1%, 0 05-1%, 0.5-2%, 1-5%, 2.5-7.5%, 5-10%, 7.5-12.5%, 10-15%,12.5-17.5%, 15-20%, 17.5-22.5%, 20-25%, 22.5-27.5%, 25-30%, 27.5-32.5%,30-35%, 32.5-37.5%, or 35-40% per weight of the second composition. Inembodiments where the second composition comprises more than onechromophore, the first chromophore may be present in an amount of about0.01-40% per weight of the second composition, and a second chromophoremay be present in an amount of about 0.0001-40% per weight of the secondcomposition. In certain embodiments, the first chromophore is present inan amount of about 0.001-0.1%, 005-1%, 0.5-2%, 3-5%, 2.5-7.5%, 5-10%,7.5-12.5%, 10-15%, 12.5-17.5%, 15-20%, 17.5-22.5%, 20-25%, 22.5-27.5%,25-30%, 27.5-32.5%. 30-35%, 32.5-37.5%, or 35-40% per weight of thesecond composition. In certain embodiments, the second chromophore ispresent in an amount of about 0.001-0.1%, 0.05-1%, 0.5-2%, 1-5%,2.5-7.5%, 5-10%, 7.5-12.5%, 10-15%, 12.5-17.5%, 15-20%, 17.5-22.5%,20-25%, 22 5-27.5%, 25-30%. 27.5-32.5%, 30-35%, 32.5-37.5%, or 35-40%per weight of the second composition. In certain embodiments, the amountof chromophore or combination of chromophores may be in the amount ofabout 0.05-40.05% per weight of the second composition. In certainembodiments, the amount of chromophore or combination of chromophoresmay be in the amount of about 0.001-0,1%, 0.05-1%, 0.5-2%, 1-5%,2.5-7.5%, 5-10%, 7.5-12.5%, 10-15%, 12.5-17.5%, 15-20%, 17.5-22.5%,20-25%, 22.5-27.5%, 25-30%, 27.5-32.5%, 30-35%, 32.5-37.5%, or 35-40.05%per weight of the second chromophore.

In some other embodiments, the first composition may comprise the firstchromophore in a liquid or as a powder, and the second composition maycomprise a gelling composition for thickening the first composition. Theoxygen-releasing agent may be contained in the second composition or ina third composition in the kit. In some embodiments, the kit includescontainers comprising the compositions of the present disclosure. Insome embodiments, the kit includes a first container comprising a firstcomposition that includes the oxygen-releasing agent, and a secondcontainer comprising a second composition that includes at least onechromophore. The containers may be light impermeable, air-tight and/orleak resistant. Exemplary containers include, but are not linuited to,syringes, vials, or pouches. The first and second compositions may beincluded within the same container but separated from one another untila user mixes the compositions. For example, the container may be adual-chamber syringe where the contents of the chambers mix on expulsionof the compositions from the chambers. In another example, the pouch mayinclude two chambers separated by a frangible membrane. In anotherexample, one component may be contained in a syringe and injectable intoa container comprising the second component.

The biophotonic composition may also be provided in a containercomprising one or more chambers for holding one or more components ofthe biophotonic composition, and an outlet in communication with the oneor more chambers for discharging the biophotonic composition from thecontamer. In one embodiment, discharging the biophotonic compositionscauses the components of the composition to mix to form a biophotoniccomposition which has less than 1.5% leaching properties.

In other embodiments, the kit comprises a systemic or topical drug foraugmenting the treatment of the composition. For example, the kit mayinclude a systemic or topical antibiotic or hormone treatment for acnetreatment or wound healing.

Written instructions on how to use the biophotonic composition inaccordance with the present disclosure may be included in the kit, ormay be included on or associated with the containers comprising thecompositions of the present disclosure.

In certain embodiments, the kit may comprise a further component whichis a dressing. The dressing may be a porous or semi-porous structure forreceiving the biophotonic composition. The dressing may comprise wovenor non-woven fibrous materials.

In certain embodiments of the kit, the kit may further comprise a lightsource such as a portable light with a wavelength appropriate toactivate the chromophore in the biophotonic composition. The portablelight may be battery operated or re-chargeable.

In certain embodiments, the kit may further comprise one or morewaveguides.

Identification of equivalent compositions, methods and kits are wellwithin the skill of the ordinary practitioner and would require no morethan routine experimentation, in light of the teachings of the presentdisclosure. Practice of the disclosure will be still more fullyunderstood from the following examples, which are presented herein forillustration only and should not be construed as limiting the disclosurein any way.

EXAMPLES

The examples below are given so as to illustrate the practice of variousembodiments of the present disclosure. They are not intended to limit ordefine the entire scope of this disclosure.

Example 1

The photodynamic properties of (i) Fluorescein sodium salt at about 0.09mg/mL, (ii) Eosin Y at about 0.305 mg/mL, and (iii) a mixture ofFluorescein sodium salt at about 0.09 mg/mL and Eosin Y at about 0.305mg/mL in a gel according to an embodiment of the present disclosure(comprising about 12% carbamide peroxide), were evaluated. A flexstation384 II spectrometer was used with the following parameters: modefluorescence, excitation 460 nm, emission spectra 465-750 nm. Theabsorption and emission spectra are shown in FIGS. 6 a and 6 b whichindicate an energy transfer between the chromophores in the combination.

Example 2

The photodynamic properties of (i) Fluorescein sodium salt at 0.18 mg/mLfinal concentration, (ii) Eosin Y at about 0.305 mg/mL, and (iii) amixture of Fluorescein sodium salt at about 0.18 mg/mL and Eosin Y atabout 0.305 mg/mL in an aqueous solution were evaluated. A flexstation384 II spectrometer was used with the following parameters: modefluorescence, excitation 460 nm, emission spectra 465-750 nm. Theabsorption and emission spectra are shown in FIGS. 7 a and 7b whichindicate an energy transfer between the chromophores in the combination.

Example 3

The photodynamic properties of (i) Rose Bengal at about 0.085 mg/mL,(ii) Fluorescein sodium salt at about 0.44 mg/mL final concentration,(ii) Eosin Y at about 0.305 mg/mL, and (iii) a mixture of (i), (ii) and(iii) in a gel comprising about 12% carbamide peroxide (Set A),according to an embodiment of the invention, were evaluated. Aflexstation 384 II spectrometer was used with the following parameters:mode fluorescence, excitation 460 nm, enussion spectra 465-750 nm. Theabsorbance and emission spectra are shown in FIGS. 8 a and 8 b whichindicate an energy transfer between the chromophores in the chromophorecombination.

Example 4

The photodynamic properties of (i) Rose Bengal at about 0.085 mg/mL,(ii) Fluorescein sodium salt at about 0.44 mg/mL final concentration,(ii) Eosin Y at about 0.305 mg/mL, and (iii) a mixture of (i), (ii) and(iii) in an aqueous solution (Set A), were evaluated. A flexstation 384II spectrometer was used with the following parameters: modefluorescence, excitation 460 nm, emission spectra 465-750 nm. Theabsorbance and emission spectra are shown in FIGS. 9 a and 9 b whichindicate an energy transfer between the chromophores in the chromophorecombination, in the absence of an oxygen-releasing agent.

Energy transfer was also seen between: Eosin Y and Rose Bengal; PhloxineB and EosinY; Phloxine B, EosinY and Fluorescein, amongst othercombinations. It is to be reasonably inferred that energy transfer canalso occur in biophotonic compositions of the present disclosure.

Example 5

A randomized, split-face clinical trial of 12 weeks was performed on 90patients (ages 14-30) having moderate to severe facial acne. Moderatefacial acne was defined as having “an Investigator's Global Assessment(IGA) of 3 with 20-40 inflammatory lesions and no more than 1 nodule”.Severe facial acne was defined as having “an IGA of 4 with more than 40inflammatory lesions with the presence of more than 2 nodules and/orpresence of sever erythema and inflammatory scarring type lesion”. Foreach patient, one randomly selected side of the face was treated twice aweek for 6 weeks with a biophotonic composition comprising Eosin Y andan oxygen-releasing agent, and exposed to light from an LED source (peakwavelength range 400-470 nm) for about 5 minutes. Other hemifaceremained untreated for the 6 week period. Both the treated and untreatedsides of the face were evaluated after 12 weeks. Results are presentedin Tables 1-5 below. The treatment was well tolerated by the patientsand there were no serious adverse events. 80% of patients completed thestudy with no adverse events reported.

At week 4, there was a 30% reduction in inflammatory lesions (includingpapules, pustules and nodules) for the treated group compared to 9.0%reduction for untreated. At week 6, the reduction was 46.8% for treatedand 18.4% for untreated, and at week 12, the reduction was 59.2% fortreated and 35.6% for untreated.

TABLE 1 Total reduction from baseline in IGA at week 12 of more than orequal to 2 grades and less than 2 grades for treated and untreatedhemifaces. Total reduction from baseline in IGA at week 12 Treated (n =89) Untreated (n = 89) ≥2 grades 46 (51.7%)* 16 (18.0%) <2 grades 43(48.3%)* 73 (82.0%) *P value < 0.0001

TABLE 2 Total reduction from baseline in IGA at week 12 of more than orequal to 1 grade and less than 1 grade for treated and untreatedhemifaces. Total reduction Total reduction from baseline from baselinein IGA at week 12 in IGA at week 6 (n = 89) Treated Untreated TreatedUntreated ≥1 grade 79 62 71 40 (88.8%)* (69.7%) (79.8%)+ (44.9%)  <1grade 10 27 18 49 (11.2%)* (30.3%) (20.2%)+ (55.1%) *P value < 0.0001 +Pvalue < 0.0001

TABLE 3 Total reduction from baseline in IGA at weeks 6 and 12 to grade0 and grade 1 and grades 2, 3 or 4 for treated and untreated hemifaces.Total reduction Total reduction from baseline in from baseline in IGA atweek 12 IGA at week 6 (n = 89) Treated Untreated Treated Untreated ToGrade 29 (32.6%)* 10 (11.2%) 16 (18.0%)+ 6 (6.7%) 0 or 1 To Grade(67.4%)* 79 (88.8%) 73 (82.0%)+ 83 (93.2%) 2, 3 or 4 *P value < 0.0001+P value < 0.0213

TABLE 4 Proportion of patients showing at least 40% reduction frombaseline in inflammatory lesion count (includes papules, pustules andnodules) at weeks 6 and 12 for treated and untreated hemifaces. Totalreduction Total reduction from baseline in from baseline in inflammatorylesion inflammatory lesion count at weck 12 count at week 6 TreatedUntreated Treated Untreated (n = 87) (n = 87) (n = 87) (n = 87) ≥40% 71(81.6%)* 40 (46.0%) 56 (64.4%)+ 27 (31.0%) <40% (18.4%)* 47 (54.0%) 31(35.6%)+ 60 (69.0%) *P value < 0.0001 +P value < 0.0001

TABLE 5 Summary of inflammatory lesion count and absolute changes byhemiface. Difference Inflam- (Treated- matory Treated UntreatedUntreated) Lesion Lesion Lesion Change Count* Count Change Count Change(Absolute) Change Base- n 90 90 90 line Mean 23.0 23.3 −0.3 (SD) (13.79)(15.41) (7.10) Week n 87   87 87 87 87 87 4 Mean 16.3  −6.9 21.2 −2.34.9 −4.6 (SD) (10.82) (6.55) (14.39) (5.44) (7.58) (7.74) Week n 87   8787 87 87 87 6 Mean 12.4 −10.9 19.0 −4.5 −6.6 −6.3 (SD) (8.35) (8.85)(13.92) (7.24) (8.34) (9.35) p <0.0001 <0.0001 value Week n 87   87 8787 87 87 12 Mean  9.5 −13.7 15.0 −8.5 −5.5 −5.2 (SD) (7.10) (11.52)(11.33) (11.04) (7.37) (9.05) p <0.0001 <0.0001 value *Includes papules,pustules and nodules

FIG. 14 is an emission spectrum showing the intensity over time of thelight being emitted from the biophotonic composition.

Example 6—Leaching Test Using Polycarbonate Membrane

FIG. 5 depicts an experimental setup of an in vitro release test forevaluating leaching of the chromophore(s) or other components (e.g.,oxygen releasing agents) from the biophotonic compositions of thepresent disclosure. In this in vitro test, a 2 mm thick layer of thebiophotonic composition is applied on the surface of a 3 cm diameterpolycarbonate (PC) membrane with pore size of 3 μm. It will beappreciated that other membranes with different pore sizes can also beused. The membrane is in direct contact with phosphate saline buffer(PBS) or PBS containing 4% bovine serum albumin (PBS/BSA) contained in aclosed compartment (i.e., the receptor compartment). The biophotoniccomposition is then illuminated with an activating light for anappropriate period of time (e.g., 5 min) at an appropriate distance(e.g. 5 cm from the light source). Samples (100 μl×2) are then takenfrom the receptor compartment at different time points (e.g., at 5, 10,20, and 30 min), and evaluated for concentration of the chromophore(s)or any other components of the biophotonic composition usingspectrophotometry or any other suitable method.

For example, when the chromophore being tested is eosin, a wavelength of517 nm (absorbance) may be used. The concentration of the chromophoremay then be calculated based on the chromophore standards of knownconcentration prepared in PBS or PBS/BSA and measured at the same time.Moreover, the presence of peroxide (i.e., an indicator of the oxygenreleasing agents) may be assessed using peroxide test sticks (e.g.Quantofix Peroxide 25, Sigma Aldrich).

Table 9 summarizes leaching data for different biophotonic compositionsaccording to the present disclosure. The amount of hydrogen peroxidefound in the receptor compartment was very low for all compositions inTable 9. The detection method of chromophore by spectrophotometry canmeasure the chromophore concentration from 0.2 μg/ml. The release ofchromophores increased overtime but was less than 15% even after 30minutes incubation which is longer than a treatment time according toembodiments of the present disclosure.

TABLE 9 Percentage of chromophores released from biophotoniccompositions according to embodiments of the present disclosure, withtime of incubation. Percentage chromophore released into receptorcompartment from composition with time of incubation (n = 3) 5 mins 10mins 20 mins 30 mins Eosin Y (0.011%) in a carbamide gel Not Not 0.750.78 (glycerine, propylene glycol, carbopol detectable detectablepolymer, urea peroxide) Fluorescein (0.2%) in a carbamide gel 2.71 4.854.72 4.84 Rose Bengal (0.2%) in a carbamide gel 2.39 3.32 5.26 5.21 RoseBengal (0.1%) + Fluorescein (0.1%) 2.91 5.21 8.48 8.43 in a carbamidegel Phloxin B (0.2%) in a carbamide gel 0.54 2.39 4.62 4.50 Eosin Y(0.2%) in a carbamide gel 2.77 2.72 6.56 9.08 Phloxin B (0.1%) andFluorescein (0.1%) 2.28 4.49 7.56 11.02 in a carbamide gel Phloxin B(0.1%) and Rose Bengal 2.41 2.36 5.14 4.90 (0.1%) in a carbamide gelEosin Y (0.1%) + Phloxin B (0.1%) in a 3.84 6.25 10.08 12.00 carbamidegel Eosin Y (0.1%) + Rose Bengal (0.1%) a 3.04 4.28 6.63 8.12 carbamidegel Eosin Y (0.1%) + Fluorescein (0.1%) in a 2.96 3.99 5.78 7.58carbamide gel Phloxin B (0.1%) + Eosin Y (0.1%) in a 1.00 2.3 4.48 5.80carbopol polymer gel Eosin Y (0.2%) in a carbopol polymer gel 6.78 8.214.38 17.89 including urea peroxide Phloxin B (0.1%) + Eosin Y (0.1%) ina 0.51 0.25 1.79 3.14 5% gelatin gel Rose Bengal (0.1%) + Eosin Y (0.1%)in 0 0.39 1.39 2.15 a 5% gelatin gel

Example 7—Angiogenic Potential of the Biophotonic Composition of theDisclosure

A human skin model was developed to assess the angiogenic potential ofthe biophotonic composition of the present disclosure. Briefly, abiophotonic composition a biophotonic composition comprising Eosin Y andErythrosine was placed on top of a human skin model containingfibroblasts and keratinocytes. The skin model and the composition wereseparated by a nylon mesh of 20 micron pore size. The composition wasthen irradiated with blue light (‘activating light’) for 5 minutes at adistance of 5 cm from the light source. The activating light consistedof light emitted from an LED lamp having an average peak wavelength ofabout 400-470 nm, and a power intensity measured at 10 cm of 7.7 J/cm²to 11.5 J/cm². Upon illumination with the activating light, thebiophotonic composition emitted fluorescent light (FIG. 4 ). Since thebiophotonic composition was m limited contact with the cells, thefibroblasts and keratinocytes were exposed mainly to the activatinglight and the fluorescent light emitted from the biophotoniccomposition. Conditioned media from the treated human 3D skin model werethen applied to human aortic endothelial cells previously plated inmatrigel. The formation of tubes by endothelial cells was observed andmonitored by microscopy after 24 hours The conditioned medium from 3Dskin models treated with light illumination induced endothelial tubeformation in vitro, suggesting an indirect effect of the light treatment(blue light and floorescence) on angiogenesis via the production offactors by fibroblasts and keratinocytes. Plain medium and conditionedmedium from untreated skin samples were used as a control, and did notinduce endothelial tube formation.

FIG. 15 is an emission spectrum showing the intensity over time of thelight being emitted from the biophotonic composition.

Example 8—Protein Secretion and Gene Expression Profiles

Wounded and unwounded 3D homan skin models (EpiDermFT, MatTekCorporation) were used to assess the potential of a biophotoniccomposition of the present disclosure to trigger distinct proteinsecretion and gene expression profiles. Briefly, a biophotoniccomposition comprising Eosin and Erythrosine were placed on top ofwounded and unwounded 3D human skin models cultured under differentconditions (with growth factors, 50% growth factors and no growthfactors). The skin models and the composition were separated by a nylonmesh of 20 micron pore size. Each skin model-composition combination wasthen irradiated with blue light (‘activating light’) for 5 minutes at adistance of 5 cm from the light source. The activating light consistedof light emitted from an LED lamp having an average peak wavelength ofabout 440-470 nm, a power density of 60-150 mW/cm2 at 5 cm, and a totalintensity after 5 minutes of about 18-39 J/cm2. The controls consistedof 3D skin models not illuminated with light.

Gene expression and protem secretion profiles were measured 24 hourspost-light exposure. Cytokine secretion was analyzed by antibody arrays(RayBio Human Cytokine antibody array), gene expression was analyzed byPCR array (PAHS-013A, SABioscience) and cytotoxicity was determined byGAPDH and LDH release. Results (Tables 1 and 2) showed that the lighttreatment is capable of increasing the level of protein secreted andgene expression involved in the early inflammatory phase of woundhealing in wounded skin inserts and in non-starvation conditions Instarvation conditions mimicking chronic wounds, there was no increase inthe level of inflammatory protein secreted when compared to the control.Interestingly, the effect of the light treatment on unwounded skinmodels has a much lower impact at the cellular level than on woundedskin insert, which suggests an effect at the cellular effect level ofthe light treatment. It seems to accelerate the inflammatory phase ofthe wound healing process. Due to the lack of other cell types such asmacrophages in the 3D skin model, the anti-inflammatory feed-back isabsent and may explain the delay in wound closure. Cytoxicity was notobserved in the light treatments.

TABLE 6 List of proteins with statistically significant differencesecretion ratio between treated and untreated control at day 3. Twoarrows mean that the ratio was over 2 folds. Medium 1X Medium 0.5XMedium 0X Increase ENA78 p = 0.04 ↑↑ Angiogenin p = 0.03 ↑ H-IR4/ST2 p =0.02 ↑↑ CXCL16 p = 0.04 ↑ MMP3 p = 0.01 ↑↑ MCP-2 p = 0.04 ↑↑ DecreaseBMP6 p = 0.01 ↓ BMP6 p = 0.02 ↓ TNFα p = 0.005 ↓

TABLE 7 List of genes with statistically significant differenceexpression ratio between treated and untreated control during the first24 hours. Two arrows mean that the ratio was over 2 folds. Medium 1XMedium 0.5X Medium 0X Increase CTGF p = 0.02 ↑ CTGF P = 0.04 ↑ MMP3 p =0.007 ↑↑ ITGB3 p = 0.03 ↑ ITGB3 p = 0.05 ↑ LAMA1 p = 0.03 ↑ MMP1 p =0.03 ↑ MMP1 p = 0.02 ↑↑ ITGA2 p = 0.03 ↑ MMP3 p = 0.01 ↑ MMP10 p = 0.003↑↑ THBS1 P = 0.02 ↑ MMP3 p = 0.007 ↑↑ MMP8 p = 0.02 ↑↑ THBS1 p = 0.03 ↑Decrease HAS1 p = 0.009 ↓↓ NCAM1 p = 0.02 ↓↓ NCAM1 p = 0.05 ↓↓ VCAN p =0.02 ↓ VCAM1 p = 0.03 ↓↓ LAMC1 p = 0.002 ↓ COL7A1 p = 0.04 ↓ COL6A1 p =0.007 ↓ CTNNA1 p = 0.03 ↓ MMP7 p = 0.003 ↓

Example 9—Collagen Formation in Skin

A randomized, placebo-controlled, single-blinded, split face and singlehand study of 32 patients, split into 4 groups (A, B, C and D), assessedthe safety and efficacy of treatment once a week for 4 weeks: (A) “lightalone”—light, according to an embodiment of the present disclosure,comprising light from an LED source having an average peak wavelength ofabout 400-490 nm at a power density of less than 150 mW/cm² for 5minutes; and a placebo formulation; (B) “light+gel”—light as in (A) plusbiophotonic gel according to an embodiment of the present disclosure);(C) “gel alone”—biophotonic gel as in (B) and a sham light (white LEDlight); and (D) 0.1% retinoic based cream. Skin biopsies were obtainedbefore treatment and 12 weeks after treatment from the treatment site.Histological samples of the skin biopsies were graded by an independentand experienced pathologist blinded to the treatment assignment. Theresults are presented in Table 8 below and show that the light treatmentwith and without the biophotonic gel, according to embodiments of thepresent disclosure, showed a 287% and 400% increase from the baseline,respectively, in collagen clusters as viewed through Gomori Trichomestaining, in the treated areas of skin. There were no serious adverseevents. There was no reported or observed photosensitivity, inflammationor pain.

TABLE 8 Semi-quantitative histological collagen evaluation Treatment %increase in collagen Photoactivatable composition excited with light 400having 460 nm peak wavelength Placebo composition + light having 460 nmpeak 287 wavelength Retinol cream with no light 189 Placebo compositionwith white light 150

Example 10—Flap Closure

A caudally based rectangular flap was elevated in the back of Wistarrats. A silicone sheet was inserted beneath the skin flap to preventadhesion and reperfusion of the flap from the underlying tissues.Following flap closure, a biophotonic gel according to an embodiment ofthe present disclosure (including chromophores and hydroscopic agents)was applied onto the dorsal flap in a thin monolayer (2 mm) and exposedto a light, for 5 minutes, from a LED light source having a peakwavelength of about 440-470 nm and a bandwidth of about 18-23 nm. Thebiophotonic gel was removed and skin specimens were collected fromdifferent areas in the flap for histological analyses nine dayspost-treatment. The treated group demonstrated a significantly greaternumber of Ki67-positive-staining events (P=0.02) compared to those inthe non-treated group these results, suggesting that the treatment maymodulate the proliferation of the cells involved in wound healing (FIG.11 ). Following examination by an external pathologist, the treatmentgroup was associated with a significant (P<0.05) decrease in thecoagulative necrosis in the epidermis and an increase of the fibrillarstroma (dermis) as compared to the control group.

Example 11—Evaluation of Removal of Biophotonic Composition from EthanolSoaked Paper

Regular white print paper was soaked in 70% ethanol (EtOH). A 2 mmthickness of different embodiment's of biophotonic compositionsaccording to the present disclosure (Table 10) were placed onto thesoaked paper and left for 5 minutes. After 5 minutes, the compositionswere washed off with 70% EtOH. A composition comprising Eosin (0.017%),silica particles, modified starch, and hydrogen peroxide was alsotested.

The results show that biophotonic compositions of the present disclosureincluding a carbamide gel do not stain white paper. A compositioncontaining Eosin and another hydrophilic polymer (starch) in combinationwith silica particles did stain the paper.

TABLE 10 Evaluation of removal of biophotonic composition from paperColour of paper Biophotonic composition after washing Eosin (0.017%),silica particles, modified Orange/red stain on starch, hydrogen peroxide(included for paper observed. comparison only). Eosin (0.011%) in a ureaperoxide, glycerin, Substantially white - propylene glycol, carbopol,hyaluronic acid, no staining observed. glucosamine gel. Eosin (0.011%) +carbamide peroxide + 1.8% Substantially white - carbopol 940 no stainingobserved.

Example 12—Evaluation of Heat Dissipation During Illumination of aBiophotonic Composition

A 3 mm thick layer of a biophotonic composition according to anembodiment of the present disclosure comprising a fluorescentchromophore in a gel according to an embodiment of the presentdisclosure was applied on the skin of hands of volunteers with differentskin types and illuminated for 5 minutes with a blue LED light having apower density of about 50 to 150 mW/cm² at a distance of 5 cm from thelight. A thermometer probe was placed within the composition, at thesurface of the skin, and the temperature was monitored in real-timeduring illumination of the composition. The skin temperature with nocomposition but the same light illumination was also measured for thesame volunteers. The skin types tested were, according to Fitzpatrickclassification scales, type III (white skin, sometimes burns andgradually tans), type IV (beige to brown skin, rarely burns and easilytans) and type VI (black skin, never burns, very easily tans). Theresults are shown in table 7.

TABLE 11 Temperature of skin under biophotonic composition duringillumination for 5 minutes compared to temperature skin with nocomposition and illumination alone Minimum-maximum Minimum-maximumtemperature of skin under temperature of skin composition during 5 minswithout composition during of illumination/° C. 5 mins. ofillumination/° C. (Average over 5 mins/° C.) (Average over 5 mins/° C.)Skin Type II 26.5-35.1 (32.2) 28.7-39.1 (36.2) Skin Type IV 27.6-39.9(36.1) 31.4-39.9 (37.0) Skin Type VI 28.5-39.9 (35.6) 29.6-40.0 (37.4)

All skin types with biophotonic composition applied demonstrated aslower temperature increase compared to bare skin (no biophotoniccomposition), and so the biophotonic composition conferred a buffereffect. After 5 minutes of light illumination, the temperature of theskin under the biophotonic composition for all volunteers reached amaximum of 39.9° C., compared to 40° C. with light alone and bare skin.Overall no pain, burning or discomfort was felt by the volunteers.

Example 13—Selecting the Concentration of Chromophore in the BrophotonicComposition

The fluorescence spectra of biophotonic compositions with differentconcentrations of chromophores were investigated using aspectrophotometer and an activating blue light. Exemplary fluorescencespectra of Eosin Y and Fluorescein are presented in FIG. 12 . It wasfound that emitted fluorescence from the chromophore increases rapidlywith increasing concentration but slows down to a plateau with furtherconcentration increase. Activating light passing through the compositiondecreases with increasing chromophore composition as more ts absorbed bythe chromophores. Therefore, the concentration of chromophores inbiophotonic compositions of the present disclosure can be selectedaccording to a required ratio and level of activating light andfluorescence treating the tissue based on this example. In someembodiments, it will be after the zone of rapid increase, i.e. between0.5 and 1 mg/mL for Eosin Y (FIG. 12 ).

Therefore, concentration can be selected according to requiredactivating light and fluorescence. In some embodiments, it will be afterzone of rapid increase, i.e. between 0.5 and 1 mg/mL for Eosin Y (FIG.12 ).

Example 14—Eosin and Rose Bengal Act in a Synergistic Manner

The synergy between two chromophores according to various embodiments ofthe present disclosure was investigated by preparing the following:

1—Eosin Y (0.035%)+Rose Bengal (0 085%) in a 12% carbamide gel)2—Rose Bengal (0.085%) in a 12% carbamide gel

Rose Bengal is known to have a high quantum yield in terms of oxygenproduction in the presence of oxygen-releasing agents whenphotoactivated by green light. Eosin Y is known to have a high quantumyield in terms of emitted fluorescent light when photoactivated and canbe at least partially activated by blue light when in a gel.Photoactivated Eosin Y does not have a high quantum yield in terms ofoxygen production in the presence of oxygen-releasing agents. When EosinY and Rose Bengal are combined, it appears that both chromophores areactivated by the same blue light as evidenced by FIG. 13 .

FIG. 13 , left panel, shows a photograph of the composition when viewedunder a light microscope (×250) before exposure to an activating light.Very few bubbles were seen in both compositions. Following illuminationwith blue light a dramatic increase in bubbles was seen with thecomposition comprising a combination of Eosin Y and Rose Bengal, but notwith the composition comprising Rose Bengal alone. This suggests thatthere is a transfer of energy from Eosin Y to Rose Bengal leading to theform oxygen species.

1.-26. (canceled)
 27. A biophotonic composition comprising: a firstchromophore; and a gelling agent present in an amount sufficient to gelthe composition and render the biophotonic composition substantiallyresistant to leaching such that less than 15% by weight of the totalchromophore amount leaches out of the biophotonic composition in use.28. The biophotonic composition of claim 27, wherein the substantialresistance to leaching comprises less than 15% of the total chromophoreamount leaching out of the biophotonic composition into a phosphatesaline buffer solution contained in a receptor compartment, through a2.4-3 cm diameter polycarbonate (PC) membrane with a thickness of 10microns and a pore size of 3 microns, having a top side onto which a 2mm thick layer of the biophotonic composition is placed for 5 minutes atroom temperature and pressure, and a bottom side which is in directcontact with the phosphate saline buffer solution.
 29. The biophotoniccomposition of claim 27, wherein the biophotonic composition istranslucent.
 30. The biophotonic composition of claim 29, wherein thetranslucency comprises at least 20%, 30%, 40%, 45%, 50%, 55%, 60%, 65%,70%, 75%, 85%, 90%, 95% or 100% transmittance of light through a 2 mmthick composition.
 31. The biophotonic composition of claim 27, whereinthe gelling agent is a synthetic polymer selected from the groupconsisting of vinyl polymers, polyoxyethylene-polyoxypropylenecopolymers, poly(ethylene oxide), acrylamide polymers and derivatives orsalts thereof.
 32. The biophotonic composition of claim 27, furthercomprising a humectant.
 33. The biophotonic composition of claim 32,wherein the humectant is glycerine.
 34. The biophotonic composition ofclaim 27, further comprising an oxygen-releasing agent.
 35. Thebiophotonic composition of claim 34, wherein the oxygen-releasing agentis selected from the group consisting of hydrogen peroxide, carbamideperoxide, benzoyl peroxide, peroxy acid, alkali metal peroxides, alkalimetal percarbonates, peroxyacetic acid, and alkali metal perborates. 36.The biophotonic composition of claim 27, wherein the first chromophoreis selected from the group consisting of Eosin Y, Eosin B, Erythrosin B,Fluorescein, Rose Bengal and Phloxin B.
 37. The biophotonic compositionof claim 27, wherein the first chromophore is present in an amount ofabout 0.001% to about 40% by weight of the total composition, preferablyabout 0.005% to about 2% by weight of the total composition.
 38. Thebiophotonic composition of claim 27, wherein the biophotonic compositionfurther comprises a second chromophore.
 39. The biophotonic compositionof claim 27, wherein the first chromophore has an emission spectrum thatoverlaps at least 20% with an absorption spectrum of the secondchromophore.
 40. The biophotonic composition of claim 27, wherein thefirst chromophore is Eosin Y, and the second chromophore is one or moreof Fluorescein, Phloxine B and Erythrosine B.
 41. The biophotoniccomposition of claim 27, wherein the first chromophore is Fluorescein,and the second chromophore is Eosin Y.
 42. A biophotonic composition,comprising a first chromophore in a carrier medium, wherein thebiophotonic composition is encapsulated in a membrane which membranelimits leaching of the first chromophore such that less than 15% of thetotal chromophore amount leaches out of the biophotonic composition inuse.
 43. The biophotonic composition of claim 49, wherein the membraneis translucent.
 44. The biophotonic composition of claim 42, wherein themembrane is selected from the group consisting of a lipid, a polymer,gelatin, cellulose, and cyclodextrins.
 45. The biophotonic compositionof claim 42, wherein the composition further comprises a dendrimer. 46.A method for providing cosmetic treatment, comprising: applyingtopically the biophotonic composition according to claim 27 to a skin;and illuminating said biophotonic composition with light having awavelength that overlaps with an absorption spectrum of the firstchromophore.